US20150030615A1

US20150030615A1 - Biomarkers for cancer stem cells and related methods of use

Info

Publication number: US20150030615A1
Application number: US14/384,026
Authority: US
Inventors: Alan G. Derr; David T. Weaver; Irina Shapiro; Daniel W. Paterson; Jonathan A. Pachter
Original assignee: Verastem Inc
Current assignee: Verastem Inc
Priority date: 2012-03-09
Filing date: 2013-03-08
Publication date: 2015-01-29
Also published as: WO2013134649A1

Abstract

Novel methods of classifying subjects as candidates for treatment with a cancer associated mesenchymal cell, tumor initiating cancer cell, or cancer stem cell inhibitor treatment and subsequent administration of the cancer associated mesenchymal cell, tumor initiating cancer cell, or cancer stem cell inhibitor are disclosed within.

Description

CROSS-REFERENCE TO RELATED APPLICATIONS

This application claims the priority of U.S. provisional application Ser. No. 61/609,031, filed Mar. 9, 2012, which is incorporated by reference herein in its entirety.

BACKGROUND

Currently available therapeutic regimens are ineffective in treating many cancers. Cancer stem cells (CSCs), cancer associated mesenchymal cells, or tumor initiating cancer cells, comprise a unique subpopulation of a tumor and have been identified in a large variety of cancer types. Although this subpopulation of cells constitutes only a small fraction of a tumor, they are thought to be the main cancer cells responsible for tumor initiation, growth, and recurrence. Given that current cancer treatments have, in large part, been designed to target rapidly proliferating cells, this subpopulation of cells, which is often slow growing, may be relatively more resistant to these treatments. Therefore, methods to identify cancer patients likely to respond positively to a treatment comprising an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells are needed; and can provide the basis for subsequent administration of a treatment comprising an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells; to this candidate group of cancer patients.

SUMMARY OF INVENTION

The present invention provides a method for classifying subjects likely to respond to a particular therapeutic regimen for treating cancer. The method is based, at least in part, on the characterization of signals (e.g., the level of gene expression) possessed by a candidate subject population for treatment with a preselected drug. In general, the method involves identifying differences in candidate and non-candidate subject populations, where for example, a subject population has a gene expression profile associated with a candidate or non-candidate classification. The method can further comprise administration of the therapeutic regimen to the candidate population based on the characterized gene expression profile.
In an aspect, the invention features a method of evaluating or treating a subject, comprising: (a) optionally, acquiring a subject sample, e.g., a tissue sample, such as a biopsy; bodily fluids, such as blood or plasma (b) acquiring a value or values that is a function of the level of gene expression for each of a plurality of genes selected from a first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or seventh and/or eighth and/or ninth and/or tenth and/or eleventh and/or twelfth and/or thirteenth and/or fourteenth set of genes; (c) and responsive to said value or values (i) classifying the subject, e.g., classifying the subject as a candidate or non-candidate for treatment with a preselected drug, and/or treating, or withholding treatment from, the subject with a preselected drug; or (ii) administering a treatment comprising an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells, to the subject; provided that, if (c)(ii) is not performed the acquisition in (a) or (b) comprises directly acquiring; thereby evaluating or treating the subject.
In an embodiment, (a) optionally, acquiring a subject sample, e.g., a tissue sample, such as a biopsy; bodily fluids, such as blood or plasma, is performed. In an embodiment, (b) acquiring a value or values that is a function of the level of gene expression for each of a plurality of genes selected from a first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or seventh and/or eighth and/or ninth and/or tenth and/or eleventh and/or twelfth and/or thirteenth and/or fourteenth set of genes, is performed. In an embodiment, (a) optionally, acquiring a subject sample, e.g., a tissue sample, such as a biopsy; bodily fluids, such as blood or plasma, is performed; and (b) acquiring a value or values that is a function of the level of gene expression for each of a plurality of genes selected from a first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or seventh and/or eighth and/or ninth and/or tenth and/or eleventh and/or twelfth and/or thirteenth and/or fourteenth set of genes, is performed are performed.
In an embodiment, (c) responsive to said value or values (i) classifying the subject, e.g., classifying the subject as a candidate or non-candidate for treatment with a preselected drug, and/or treating, or withholding treatment from, the subject with a preselected drug; is performed. In an embodiment, (c) responsive to said value or values (ii) administering a treatment comprising an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells, to the subject; is performed. In an embodiment, (c) responsive to said value or values (i) classifying the subject, e.g., classifying the subject as a candidate or non-candidate for treatment with a preselected drug, and/or treating, or withholding treatment from, the subject with a preselected drug; is performed and (c) responsive to said value or values (ii) administering a treatment comprising an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells, to the subject; are performed.
In an embodiment, the invention features, responsive to said value or values, classifying the subject, e.g., classifying the subject as a candidate or non-candidate for treatment with a preselected drug, and/or treating, or withholding treatment from, the subject with a preselected drug, wherein the subject sample is directly acquired; thereby evaluating the subject.
In an embodiment, the invention features, responsive to said value or values, classifying the subject, e.g., classifying the subject as a candidate or non-candidate for treatment with a preselected drug, and/or treating, or withholding treatment from, the subject with a preselected drug, wherein said value or values is directly acquired; thereby evaluating the subject.
In an embodiment, the invention features, responsive to said value or values, classifying the subject, e.g., classifying the subject as a candidate or non-candidate for treatment with a preselected drug, and/or treating, or withholding treatment from, the subject with a preselected drug, wherein the subject sample and said value or values are directly acquired; thereby evaluating the subject.
In an embodiment, the invention features, responsive to said value or values, administering a treatment comprising an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells to said subject.
In an embodiment, the invention features, responsive to said value or values, classifying the subject, e.g., classifying the subject as a candidate or non-candidate for treatment with a preselected drug, and/or treating, or withholding treatment from, the subject with a preselected drug; and administering a treatment comprising an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells to said subject.
In an embodiment, the first set of genes are related to the delineation of a cancer stem cell, cancer associated mesenchymal cell, or a tumor initiating cancer cell phenotype and the determination of the attributes of the epithelial to mesenchymal transition (EMT) in cancers. For example,
(i) said first set of genes comprises or consists of:

- KLF13, ANTXR2, IFNAR1, LIX1L, CHST11, AKAP2, PALM2-AKAPT, LOC80298, C7orf2, NUCKS, DKFZP566D1346, LOC388279, FLJ31795, 6orf107, FLJ12439, ENPP2, CITED2, ZEB1, NID2, SEMA5A, DAB2, KCNMA1, PTX3, PCDH9, CREB3L1, BGN, ROR1, DLC1, GNG11, CDH11, NR2F1, PRR16, MYL9, DOCK10, LRIG1, IER3, EML1, NEBL, RGL1, MLPH, DNAJB4, FBLN5, RGS4, HAS2, ITGBL1, IGFBP4, DPT, PCOLCE, GREM1, PPAP2B, CDH2, PMP22, LUM, CHN1, CYP1B1, MME, WNT5A, POSTN, MMP2, CTGF, FBLN1, IGFBP3, DCN, PRRX1, ANXA6, PVRL3, MAP1B, TNFAIP6, CYBRD1, FBN1, NID1, OLFML3, SNAI1, SNAI2, TAGLN, SYNC, TUBA1A, PPM1D, TWIST1, FN1, TGFBR3, SERPINF1, UGDH, SRGN, FAP, PTGER4, PRKCA, FSTL1, MMP1, NRP1, FILIP1L, SCCPDH, LTBP2, XYLT1, HS3ST2, SYT11, TSHZ1, THY1, Sep9, S100A4, TNS3, ENOX1, TGFB1I1, ZEB2, LMCD1, PDGFC, ECM1, TFPI, TBX3, DDR2, PTGER2, CDKN2C, VCAN, CD44, STARD13, SNED1, ZBTB38, SDC2, TPM1, COPZ2, STC1, CDH1, KRT5, KRT6B, CD24, EPCAM, ESRP1, CLDN3, CLDN4, ERBB3, SPOCK1, FERMT2, GLYR1, LTBP1, FADS2, KANK2, PTGFR, COL11A2, KLK3, EIF2C2, ZFP41, FAM49B, PSORS1C2, MRPL42, MRPL54, MRPL47, MRPS23, EIF3S9, ALG5, DNAJC19, TPRXL, NOTCH2, RBM15, ST3GAL3, NFYA, PCNX, FBXO21, WWOX, CAMK2B, PNPLA2, CLIC5, UGCGL1, FBXL18, ADRBK1, SLC38A2, IL8RA, TAS2R14, CD300LB, GIPC3, RAD51L1, EPPK1, COL1A1, MMP9, SERPINE1, SPARC, TGFB1, TGFB3, TGFBI, TGFBR1, TGIF, TGIF2, THBS1, ANXA5, ACTG1, ARF3, ATP1B3, BAT3, CALD1, CENTD2, CLIC1, CTBS, DPYSL3, DVL3, EXT1, FGFR1, FTL, GNB2L1, GPRC5A, H2AFZ, HIF1A, IL13RA1, KDELR2, LARP1, LPIN2, MARS, MMP10, MMP14, MT2A, MYO10, NUP62, PFKFB3, PLOD2, PSMB7, PSMD8, RIN2, RYBP, SDF4, SETD5, SPP1, STAU1, TUBB3, UBE2S, XPNPEP1, CDKN1A, CHRD, H19, ID3, ID4, IGFBP7, LRP1, MSX1, NOTCH3, PROCR, GBX2, KI67, CCNB1, BUB1, KNTC2, USP22, HCFC1, RNF2, ANK3, FGFR2, CES1, COL1A2, COL3A1, COL5A2, COL6A1, ANKRD25, C10ORF56, C5ORF13, KRT81, NPAC, PLEKHC1, SEPT9, SYNC1, MBP, ABLIM1, ALDH1A3, ALOX15B, ANXA3, AP1M2, ARTN, CA2, CA9, CDH3, CDS1, COL17A1, CORO1A, CTSL2, CXADR, CYP27B1, DSC2, DSG3, DST, EPB41L4B, FGFBP1, FGFR3, FST, GJB3, GRHL2, HBEGF, HOOK1, IL18, IL1B, IRF6, ITGB4, JAG2, KLK10, KLK5, KLK7, KLK8, KRT15, KRT16, KRT17, LEPREL1, MYO5C, NDRG1, NMU, PI3, RAB25, RLN2, RNF128, S100A14, S100A7, S100A8, SERPINB1, SERPINB2, SLC2A9, SLPI, SNX10, SPINT2, SPRR1A, SPRR1B, ST14, SYK, TMEM40, TRIM29, TSPAN1, UCHL1, VSNL1, ZBED2, AQP3, CBARA1, CLCA2, CNTNAP2, CSTA, DSC3, ESRP2, FXYD3, GJB5, GPR87, IL1A, LAMA3, LAMB3, MAP7, MAPK13, MPZL2, NRG1, SCNN1A, SERPINB13, SLC7A5, SPINT1, ST6GALNAC2, TGFA, TP63, TPD52L1, VGLL1, SLC6A8, ABCA12, ANXA8, ANXA8L1, ARHGAP25, ARHGAP8, BDKRB2, BIK, C10ORF10, C10ORF116, C1ORF116, C20ORF19, C6ORF105, CELSR2, CKMT1A, CKMT1B, CLDN1, CST6, CYP4F11, E2F5, ELMO3, EPHA1, EVA1, EXPH5, F11R, FAT2, FLJ12684, FLJ20366, FZD3, GLS2, GNAL, IFI30, IGFBP2, IL1RN, IL4R, IRX4, JUP, KCNK1, KIAA0040, KIAA0888, KIAA1815, KRT14, KRT18, LAD1, LGALS7, LOC553158, LOC653562, LOC728113, LOC729884, LRRC1, LSR, MST1R, MYO1D, NAIP, OCLN, NEFM, NUP62CL, PERP, PLS1, POLR3G, PRKCH, PRRG4, PRSS8, PTPN3, RBM35A, RBM35B, RHBDF2, RHOD, RPS6KA1, RTEL1, SAA1, SAA1, SAA2, SH2D3A, SLC6A10P, SMPDL3B, SNCA, STAC, STAP2, TACSTD1, THBD, TMEM30B, TMPRSS11E, TMPRSS4, TNFRSF6B, TP73L, WWC1, XDH, ZNF165, AXL, GPR30, PAX2, PI3KA, CD133, ALDH1, BMI1, KRT19, MSI1, JAG1, p38, ESA1, FOXC2, FOXC1, TWIST2, GSC, TCF3, MLPH1, ARHGEF11a, DPF2, CASP8, BCL2, SCGN, SWAP70, KIAA0276, C10orf9, C10orf7, ALKBH, TOB2, XPR1, CD59, LRP2, PLP2, MAPK14, CXCL2, MMP7, MGP, MLF1, FLNB, SCNM1, HSPC163, C5orf18, MGC4399, CDW92, TMC4, ZDHHC2, TICAM2, KDELR3, GNPDA1, THEM2, DBR1, FLJ90709, FLJ10774, C16orf33, GAPD, LDHA, MR-1, LARS, GTPBP1, PRSS16, WFDC2, AIM1, DHRS6, DHRS4, GC15429, MGC45840, ECHDC2, GOLGIN-67, AFURS1, KIAA0436, CYP4V2, JTV1, ICMT, DNMT3A, HNMT, METTL7A, METTL2, VIL2, TPD52, ARPC5, NOL8, NSF, RAD23B, SRP54, HSPA2, PBP, THAP2, CIRBP, SNRPN, KIAA0052, DUSP10, SSR1, ERBB4, EMP1, CHPT1, LRPAP1, FLJ11752, CSTF1, KLHL20, DNAJC13, APLP2, ARGBP2, DNAJB1, NEBL, SH3BGRL, NUDT5, GABARAPL1, MAPT, DCBLD1, STK39, PAK2, CSNK2A1, PILRB, ERN1, SGKL, WEE1, MAST4, C11orf17, NUP37, CLTC, COPB2, SLC25A25, ECOP, PDE8A, STAM, TUBB, SNX6, RAB23, PLAA, STC2, LTF, ISGF3G, ATXN3, GTF3C3, GSK3B, KLF10, ELL2, ZBTB20, IRX3, ETS1, SERTAD1, MGC4251, MAFF, SFPQ, CITED4, CEBPD, EIF4E2, HS2ST1, AGPS, PGK1, ATIC, ETNK1, LG2, NCE2, MARCH8, CNOT4, RNF8, PSMA5, DPF2, AMMECR1, KIAAI287, LOC144233, LOC286505, PNAS-4, FLJ20530, HUMPD3, GC45564, CAP350, ETAA16, HAN11, DNAPTP6, C7orf25, FLJ37953, FLJ10587, C7orf36, ELP4, NDEL1, NPD014, KFZP564D172, FAM53C, IER5, LOC255783, KIAA0146, KIAA0792, LOC439994, LOC283481, CG018, LOC130576, NGFRAP1L1, KIAAI217, 4orf7, C21 orf86, C9orf64, FLJ13456, FLJ12806, FLJ39370, GATS, CCDC92, FMNL2, ARID1B, ZFHX1B, SSBP2, ARID5B, LOC157381, KPNA3, ARHGAP24, CCND2, VIM, MYCBP2, GAS7, TRAM2, BASP1, FOXO1A, POLR2A, PER1, DDIT4, CD97, BIN1, SH2B3, DDB2, EMP3, NDST2, CHST2, NT5E, PDE4A, CPS1, PTGS1, GGCX, IRF5, ZBTB16, MAP4K4, CHST7, KLF12, NFRKB, PSD, FKSG49, NIFUN, FYN, ZMYM2, CACNA1G, SLC25A16, FLII, EIF1, SEPT6, NPAC, PHF15, NUP188, ABR, CNR1, LOC283824, FSTL4, DNM1, APOBEC3G, ATP2B1, SMPD1, SLC11A1, FXYD5, C14orf139, SH3BGRL3, LUZP1, ZNF335, SH3 KBP1, MST150, PRO1073, LOC388397, FKBP5, HIPK2, DTX1, ST3GAL2, ADAMTS7, TNRC6B, CYGB, SDHAL1, LOC572558, TRIO, FRAS1, KIAA1632, POLS, EBF, MAML2, PTPRA, PLEKHG2, DYM, SOX6, ARHGEF2, ZCCHC6, PPP3CA, FAM70B, TMED5, FLJ43663, HPS1, MEF2A, ST3GAL5, SMYD3, KLF7, LOC200230, RERE, QKI, BICD1, CTNNB1, POU2F2, EIF4ENIF1, BTG1, TCHP, FLJ90709, PCTK2, KIAA1600, B7-H4, PDE4DIP, KIAA0194, HOM-TES; and
- (a) BGN, BIN1, CDH11, CDH2, CHN1, COL1A2, COL3A1, COL5A2, CREB3L1, CTGF, CYBRD1, DCN, DLC1, DNAJB4, DPT, ENPP2, FBLN1, FBLN5, FBN1, GREM1, HAS2, IGFBP3, IGFBP4, MLPH, MME, MMP2, MYL9, NEBL, NID1, NR2F1, OLFML3, PCOLCE, PMP22, POSTN, PPAP2B, PRR16, PRRX1, PTX3, PVRL3, RGL1, RGS4, ROR1, SEMA5A, TNFAIP6, TRAM2, WNT5A, ZEB1, ANXA6, CITED2, COL1A1, COL6A1, CYP1B1, DAB2, DOCK10, EML1, GNG11, IER3, ITGBL1, KCNMA1, LRIG1, LUM, MAP1B, NID2, PCDH9, PRKCA, PTGER4, PTGFR, SDC2, SERPINF1, SNED1, SYT11, TFPI, TGFBR3, VCAN, ANKRD25, C10ORF56, C5ORF13, CCDC92, CDKN2C, COPZ2, DDR2, ECM1, ENOX1, FADS2, FAP, FILIP1L, FSTL1, HS3ST2, KRT81, LMCD1, LTBP1, LTBP2, MMP1, NPAC, NRP1, PDGFC, PLEKHC1, PPM1D, PTGER2, SCCPDH, SEPT9, SPOCK1, SRGN, STARD13, STC1, SYNC1, TAGLN, TBX3, TGFB1I1, THY1, TNS3, TPM1, TSHZ1, TUBA1A, UGDH, VIM, XYLT1, ZBTB38, MBP, ABLIM1, ALDH1A3, ALOX15B, ANXA3, AP1M2, ARTN, CA2, CA9, CAMK2B, CCND2, CDH1, CDH3, CDS1, COL17A1, CORO1A, CTSL2, CXADR, CYP27B1, DSC2, DSG3, DST, EPB41L4B, FGFBP1, FGFR3, FST, GJB3, GRHL2, HBEGF, HOOK1, IL18, IL1B, IRF6, ITGB4, JAG2, KLK10, KLK5, KLK7, KLK8, KRT15, KRT16, KRT17, KRT5, KRT6B, LEPREL1, MYO5C, NDRG1, NMU, PI3, RAB25, RLN2, RNF128, S100A14, S100A7, S100A8, SERPINB1, SERPINB2, SLC2A9, SLPI, SNX10, SPINT2, SPRR1A, SPRR1B, ST14, SYK, TMEM40, TRIM29, TSPAN1, UCHL1, VSNL1, ZBED2, AQP3, CBARA1, CD24, CLCA2, CNTNAP2, CSTA, DSC3, EPCAM, ESRP2, FXYD3, GJB5, GPR87, IL1A, LAMA3, LAMB3, MAP7, MAPK13, MMP9, MPZL2, NRG1, SCNN1A, SERPINB13, SLC7A5, SPINT1, ST6GALNAC2, TGFA, TP63, TPD52L1, VGLL1, SLC6A8, ABCA12, ANK3, ANXA8, ANXA8L1, ARHGAP25, ARHGAP8, BDKRB2, BIK, C10ORF10, C10ORF116, C1ORF116, C20ORF19, C6ORF105, CELSR2, CKMT1A, CKMT1B, CLDN1, CST6, CYP4F11, E2F5, ELMO3, EPHA1, EVA1, EXPH5, F11R, FAT2, FGFR2, FLJ12684, FLJ20366, FZD3, GLS2, GNAL, IFI30, IGFBP2, IL1RN, IL4R, IRX4, JUP, KCNK1, KIAA0040, KIAA0888, KIAA1815, KRT14, KRT18, LAD1, LGALS7, LOC553158, LOC653562, LOC728113, LOC729884, LRRC1, LSR, MST1R, MYO1D, NAIP, OCLN, NEFM, NUP62CL, PERP, PLS1, POLR3G, PRKCH, PRRG4, PRSS8, PTPN3, RBM35A, RBM35B, RHBDF2, RHOD, RPS6KA1, RTEL1, SAA1, SAA1, SAA2, SH2D3A, SLC6A10P, SMPDL3B, SNCA, STAC, STAP2, TACSTD1, THBD, TMEM30B, TMPRSS11E, TMPRSS4, TNFRSF6B, TP73L, WWC1, XDH, ZNF165; and
- (b) BGN, BIN1, CDH11, CDH2, CHN1, COL1A2, COL3A1, COL5A2, CREB3L1, CTGF, CYBRD1, DCN, DLC1, DNAJB4, DPT, ENPP2, FBLN1, FBLN5, FBN1, GREM1, HAS2, IGFBP3, IGFBP4, MLPH, MME, MMP2, MYL9, NEBL, NID1, NR2F1, OLFML3, PCOLCE, PMP22, POSTN, PPAP2B, PRR16, PRRX1, PTX3, PVRL3, RGL1, RGS4, ROR1, SEMA5A, TNFAIP6, TRAM2, WNT5A, ZEB1, ANXA6, CITED2, COL1A1, COL6A1, CYP1B1, DAB2, DOCK10, EML1, GNG11, IER3, ITGBL1, KCNMA1, LRIG1, LUM, MAP1B, NID2, PCDH9, PRKCA, PTGER4, PTGFR, SDC2, SERPINF1, SNED1, SYT11, TFPI, TGFBR3, VCAN, ANKRD25, C10ORF56, C5ORF13, CCDC92, CDKN2C, COPZ2, DDR2, ECM1, ENOX1, FADS2, FAP, FILIP1L, FSTL1, HS3ST2, KRT81, LMCD1, LTBP1, LTBP2, MMP1, NPAC, NRP1, PDGFC, PLEKHC1, PPM1D, PTGER2, SCCPDH, SEPT9, SPOCK1, SRGN, STARD13, STC1, SYNC1, TAGLN, TBX3, TGFB1I1, THY1, TNS3, TPM1, TSHZ1, TUBA1A, UGDH, VIM, XYLT1, ZBTB38; and
- (c) ENPP2, CITED2, ZEB1, NID2, SEMA5A, DAB2, KCNMA1, PTX3, PCDH9, CREB3L1, BGN, ROR1, DLC1, GNG11, CDH11, NR2F1, NR2F2, PRR16, MYL9, DOCK10, LRIG1, IER3, EML1, NEBL, RGL1, MLPH, TRAM2, DNAJB4, FBLN5, RGS4, HAS2, ITGBL1, IGFBP4, BIN1, DPT, PCOLCE, GREM1, PPAP2B, CDH2, PMP22, LUM, CHN1, CYP1B1, MME, WNT5A, POSTN, MMP2, CTGF, FBLN1, IGFBP3, DCN, PRRX1, ANXA6, PVRL3, MAP1B, TNFAIP6, CYBRD1, FBN1, NID1, OLFML3, SNAI1, SNAI2, TAGLN, SYNC, TUBA1A, PPM1D, TWIST1, FN1, TGFBR3, SERPINF1, UGDH, SRGN, FAP, PTGER4, VIM, PRKCA, FSTL1, MMP1, NRP1, FILIP1L, SCCPDH, LTBP2, XYLT1, HS3ST2, SYT11, TSHZ1, THY1, SEPT9, S100A4, CCDC92, TNS3, ENOX1, TGFB1I1, ZEB2, LMCD1, PDGFC, ECM1, TFPI, TBX3, DDR2, PTGER2, CDKN2C, VCAN, CD44, STARD13, SNED1, ZBTB38, SDC2, TPM1, COPZ2, STC1, CDH1, KRT5, KRT6B, CD24, EPCAM, ESRP1, CLDN3, CLDN4, ERBB3, SPOCK1, FERMT2, GLYR1, LTBP1, FADS2, KANK2, PTGFR, ARTN, SLC7A5, EPB41L4B, KRT15, CDH3, TPD52L1, ANXA3, PI3, NDRG1, DSG3, S100A14, S100S7, KLK8, S100A8, PTX3, SPINT2, FXYD3, KRT16, RAB25, LGALS7, DSC3, COL5A2, ITGB4, TMEM40, SPRR1A, AP1M2, SERPINB13, TP73L; and
- (d) CYBRD1, ARTN, CITED2, SLC7A5, CHN1, CTGF, CSTA, MYL9, EPB41L4B, KRT15, LRIG1, CDH3, TPD52L1, ANXA3, OLFML3, CORO1A, PI3, NDRG1, DSG3, PMP22, S100A14, S100S7, KLK8, S100A8, PTX3, SPINT2, IER3, FXYD3, KRT16, TGFBR3, KRT5, RAB25, SDC2, LUM, LGALS7, KRT6B, CD24, COL1A2, GNG11, DSC3, COL5A2, RGS4, ITGB4, TMEM40, SPRR1A, AP1M2, SERPINB13, SNED1, LEPREL1, TRAM2, TP73L, and PAP2B; and
- (e) CYBRD1, ARTN, CITED2, SLC7A5, CHN1, CTGF, CSTA, MYL9, EPB41L4B, KRT15, LRIG1, CDH3, TPD52L1, ANXA3, OLFML3, CORO1A; and
- (f) PI3, NDRG1, DSG3, PMP22, S100A14, S100S7, KLK8, S100A8, PTX3, SPINT2, IER3, FXYD3, KRT16, TGFBR3; and
- (g) KRT5, RAB25, SDC2, LUM, LGALS7, KRT6B, CD24, COL1A2, GNG11, DSG3, COL5A2; and
- (h) RGS4, ITGB4, TMEM40, SPRR1A, AP1M2, AERPINB13, SNED1, LEPREL1, TRAM2, TP73L, and PAB2B; and
- (i) CYBRD1, ARTN, KRT15, ITGB4, RGS4, SPRR1B, SLC7A5, ANXA3, IL18, AP1M2, CHN1, OLFML3, ZBED2, LEPREL1, ABLIM1, CDH3, DNAJB4, TRAM2, UCHL1, CTGF; and
  - (i) CYBRD1, ARTN, KRT15, ITGB4, RGS4, SPRR1B, SLC7A5, ANXA3, IL18, AP1M2; and
  - (ii) CYBRD1, ARTN, KRT15, ITGB4, RGS4; and
- (j) CYBRD1, RGS4, CDH3, CHN1, SLC7A5, PRRX1, CITED2, COL5A2, ARTN, ZBED2, SNED1, ITGB4, LUM, PVRL3, OLFML3, BIN1, CCND2, DAB2, ANXA3, IL18; and
  - (i) CYBRD1, RGS4, CDH3, CHN1, SLC7A5, PRRX1, CITED2, COL5A2, ARTN, ZBED2; and
  - (ii) CYBRD1, RGS4, CDH3, CHN1, SLC7A5; and
- (k) CYBRD1, ARTN, RGS4, CITED2, SLC7A5, ITGB4, CHN1, PI3, TMEM40, NDRG1, SPRR1A, PCDH9, CTGF, MAP1B, DSG3, AP1M2, FBLN5, SERPINB13, PMP22, CSTA; and
  - (i) CYBRD1, ARTN, RGS4, CITED2, SLC7A5, ITGB4, CHN1, PI3, TMEM40, NDRG1; and
  - (ii) CYBRD1, ARTN, RGS4, CITED2, SLC7A5; and

(ii) said second set of genes comprises or consists of:
AXL, NOTCH3, GPR30, PROCR, PAX2, PI3KA, CD44, CD133, ALDH1, BMI1, KRT19, NRP1, MSI1, MTOR, JAG1, p38, ESA1, ZEB1, TWIST1, FOXC2, CDH2, ZEB2, FOXC1, SNAI1, SNAI2, TWIST2, GSC, TCF3, CD44, VIM, FN1, MLPH1, ARHGEF11; and
(iii) said third set of genes comprises or consists of:
At least one gene from said first set of genes and at least one of:
AXL, NOTCH3, GPR30, PROCR, PAX2, PI3KA, CD44, CD133, ALDH1, BMI1, KRT19, NRP1, MSI1, MTOR, JAG1, p38, ESA1, ZEB1, TWIST1, FOXC2, CDH2, ZEB2, FOXC1, SNAI1, SNAI2, TWIST2, GSC, TCF3, CD44, VIM, FN1, MLPH1, ARHGEF11; and
(iv) said fourth set of genes comprises or consists of:
at least two genes from said first set of genes and at least one of:
AXL, NOTCH3, GPR30, PROCR, PAX2, PI3KA, CD44, CD133, ALDH1, BMI1, KRT19, NRP1, MSI1, MTOR, JAG1, p38, ESA1, ZEB1, TWIST1, FOXC2, CDH2, ZEB2, FOXC1, SNAI1, SNAI2, TWIST2, GSC, TCF3, CD44, VIM, FN1, MLPH1, and ARHGEF11; and
(v) said fifth set of genes comprises or consists of:
beta catenin, FZD1, FZD2, FZD3, FZD4, FZD5, FZD6, FZD7, FZD8, FZD9, FZD10, LRP5, LRP6, WNT1, WNT2, WNT2B, WNT3, WNT3A, WNT4 WNT5A, WNT5B, WNT6, WNT7A, WNT7B, WNT8A, WNT8B, WNT9A, WNT9B, WNT10A, WNT10B, WNT11, WNT16, SRFP1, SRFP2, FRZB, SFRP4, SFRP5, DKK1, DKK2, DKK3, DKK4, WIF1, AKT2, PI3KA, Disheveled, Smoothened, Axin1, Axin2; and
(vi) said sixth set of genes comprises or consists of:
At least one gene from said first set of genes and at least one of:
beta catenin, FZD1, FZD2, FZD3, FZD4, FZD5, FZD6, FZD7, FZD8, FZD9, FZD10, LRP5, LRP6, WNT1, WNT2, WNT2B, WNT3, WNT3A, WNT4 WNT5A, WNT5B, WNT6, WNT7A, WNT7B, WNT8A, WNT8B, WNT9A, WNT9B, WNT10A, WNT10B, WNT11, WNT16, SRFP1, SRFP2, FRZB, SFRP4, SFRP5, DKK1, DKK2, DKK3, DKK4, WIF1, AKT2, PI3KA, Disheveled, Smoothened, Axin1, Axin2; and
(vii) said seventh set of genes comprises or consists of:
TGFbeta1, TGFbeta2, TGFbeta3, ALK1, ALK2, ALK3, ALK4, ALK5, ALK6, ALK7, TGFbetaR-II, BMPR-II, ACTR-IIA, ACTR-IIB, AMHR-II, SMAD2, SMAD3; and
(viii) said eighth set of genes comprises or consists of:
at least one gene from said first set of genes and at least one of:
TGFbeta1, TGFbeta2, TGFbeta3, ALK1, ALK2, ALK3, ALK4, ALK5, ALK6, ALK7, TGFbetaR-II, BMPR-II, ACTR-IIA, ACTR-IIB, AMHR-II, SMAD2, SMAD3; and
(ix) said ninth set of genes comprises or consists of:
S100A4, SDC2, CYP1B1, FERMT2, CYBRD1, SNAI2, CD44, BIN1, PMP22, CTGF, IGFBP3, ANXA6, MAP1B, TAGLN, ENPP2, FBLN5, PCOLCE, FN1, VIM, PPAP2B, SEM5A, VCAN, NID2, NID1, HAS2, PRRX1, TNFAIP6, CLDN4, FADS2, DDIT4, GNG11, PER1, ISCU, KPNA3, SLC44A1, FKBP5, NR2F1, MLPH, DOCK10, RGS4, IGFBP4; and
(x) said tenth set of genes comprises or consists of:
S100A4, SDC2, CYP1B1, FERMT2, CYBRD1, SNAI2, CD44, BIN1, ENPP2, FBLN5, PCOLCE, FN1, VIM, PPAP2B, SEM5A, VCAN, CLDN4, FADS2, DDIT4, GNG11, PER1, ISCU, KPNA3, SLC44A1, FKBP5, IGFBP4; and
(xi) said eleventh set of genes comprises or consists of:
SDC2, CYBRD1, SNAI2, PMP22, CTGF, IGFBP3, ANXA6, MAP1B, TAGLN, PPAP2B, HAS2, NRF2, MLPH, DOCK10, RGS4; and
(xii) said twelfth set of genes comprises or consists of:
FERMT2, BIN1, PMP22, CHN1, CTGF, IGFBP3, NID2, CLDN4, CYBRD1, PRR16, ANXA6, PCOLCE, PPAP2B, HAS2, CDH11, MLPH, DOCK10, RGS4, PVRL3, NID2, SLC44A1, FKBP5; and
(xiii) said thirteenth set of genes comprises or consists of:
FERMT2, BIN1, PMP22, CHN1, CTGF, IGFBP3, NID2, CLDN4; and
(xiv) said fourteenth set of genes comprises or consists of:
HAS2, BIN1, PCOLCE, FERMT2, CTGF, IGFBP3, NID2, SLC44A1, FKBP5, and MLPH.
In an embodiment, said plurality of genes is elected from gene sets one, i.e. (i); two i.e., (ii); three, i.e. (iii); four, i.e. (iv); five, i.e. (v); six, i.e. (vi); seven, i.e. (vii); or eight, i.e. (viii); or nine (ix); or ten (x); or eleven (xi); or twelve (xii); or thirteen (xiii); or fourteen (xiv). In an embodiment, said plurality of genes is elected from gene set (i). In an embodiment, said plurality of genes is elected from gene sets (i)a; (i)b; (i)c; (i)d; (i)e; (i)f; (i)g; (i)h; (i)i; (i)j; or (i)k. In an embodiment, said plurality of genes is elected from gene sets (i)i(i); or (i)i(ii). In an embodiment, said plurality of genes is elected from gene sets i(j)i; or (i)j(ii). In an embodiment, said plurality of genes is elected from gene sets (i)k(i); or (i)k(ii). In an embodiment, said plurality of genes is elected from gene set (ii). In an embodiment, said plurality of genes is elected from gene set (iii). In an embodiment, said plurality of genes is elected from gene set (iv). In an embodiment, said plurality of genes is elected from gene set (v). In an embodiment, said plurality of genes is elected from gene set (vi). In an embodiment, said plurality of genes is elected from gene set (vii). In an embodiment, said plurality of genes is elected from gene set (viii). In an embodiment, said plurality of genes is elected from gene set (ix). In an embodiment, said plurality of genes is elected from gene set (x). In an embodiment, said plurality of genes is elected from gene set (xi). In an embodiment, said plurality of genes is elected from gene set (xii). In an embodiment, said plurality of genes is elected from gene set (xiii). In an embodiment, said plurality of genes is elected from gene set (xiv). In an embodiment, said plurality of genes is any subset of genes in gene sets one, two, three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, or fourteen. In an embodiment, said plurality of genes is any subset of genes in gene set one. In an embodiment, said plurality of genes is any subset of genes in gene set two. In an embodiment, said plurality of genes is any subset of genes in gene set three. In an embodiment, said plurality of genes is any subset of genes in gene set four. In an embodiment, said plurality of genes is any subset of genes in gene set five. In an embodiment, said plurality of genes is any subset of genes in gene set six. In an embodiment, said plurality of genes is any subset of genes in gene set seven. In an embodiment, said plurality of genes is any subset of genes in gene set eight. In an embodiment, said plurality of genes is any subset of genes in gene set nine. In an embodiment, said plurality of genes is any subset of genes in gene set ten. In an embodiment, said plurality of genes is any subset of genes in gene set eleven. In an embodiment, said plurality of genes is any subset of genes in gene set twelve. In an embodiment, said plurality of genes is any subset of genes in gene set thirteen. In an embodiment, said plurality of genes is any subset of genes in gene set fourteen.
In an embodiment, said plurality of genes comprises at least two genes; four genes; six genes; eight genes; ten genes; twelve genes; fourteen genes; sixteen genes; eighteen genes; twenty genes; twenty five genes; thirty genes; forty genes; or any range intervening there between. In an embodiment, said plurality comprises more than forty genes.
In an embodiment, said plurality of genes comprises or consists of, a first gene and a second gene. In an embodiment, said plurality of genes further comprises, or consists of, a third gene; a third and fourth gene; a third, fourth, and fifth gene; a third, fourth, fifth, and sixth gene; a third, fourth, fifth, sixth, and seventh; a third, fourth, fifth, sixth, seventh, and eighth gene; a third, fourth, fifth, sixth, seventh, eighth and ninth gene; a third, fourth, fifth, sixth, seventh, eighth, ninth, and tenth gene. In an embodiment, said plurality of genes comprises of more than ten genes.
In an embodiment, said value or values is a function of the level of gene expression of a first gene and the level of gene expression of a second gene. In an embodiment, said value or values is a function of the level of gene expression of said first and second gene, and a third gene; a third and fourth gene; a third, fourth, and fifth gene; a third, fourth, fifth, and sixth gene; a third, fourth, fifth, sixth, and seventh gene; a third, fourth, fifth, sixth, seventh, and eighth gene; a third, fourth, fifth, sixth, seventh, eighth and ninth gene; a third, fourth, fifth, sixth, seventh, eighth, ninth, and tenth gene. In an embodiment, said value or values is a function of the level of gene expression of more than ten genes.
In an embodiment, a first value that is a function of the level of gene expression of said first gene and a second value that is a function of the level of gene expression of said second gene are acquired. In an embodiment, a first value that is a function of the level of gene expression of said first gene, a second value that is a function of the level of gene expression of said second gene, a third value that is a function of the level of gene expression of said third gene, a fourth value that is a function of the level of gene expression of said fourth gene, a fifth value that is a function of the level of gene expression of said fifth gene, a sixth value that is a function of the level of gene expression of said sixth gene, a seventh value that is a function of the level of gene expression of said seventh gene, an eighth value that is a function of the level of gene expression of said eighth gene, a ninth value that is a function of the level of gene expression of said ninth gene, and a tenth value that is a function of the level of gene expression of said tenth gene is acquired. In an embodiment, a plurality of values that is each a function of the level of gene expression of each of a plurality of genes is acquired. In an embodiment, more than ten values that is each a function of the level of gene expression of each of a plurality of genes is acquired.
In an embodiment, a first value that is a function of the level of gene expression of two or more genes of a plurality of genes and a second value that is a function of the level of gene expression of one of the genes of the plurality are acquired. In an embodiment, the invention further features the acquisition of a value or values that is a function of the level of gene expression of a gene not in said first, second, third, fourth, fifth, sixth, seventh, eighth, ninth, tenth, eleventh, twelfth, thirteenth, or fourteenth gene sets. In an embodiment, the invention further features the acquisition of a plurality of value or values that is a function of the level of gene expression of a plurality of genes not in said first, second, third, fourth, fifth, sixth, seventh, eighth, ninth, tenth, eleventh, twelfth, thirteenth, or fourteenth gene sets.
In an embodiment, the invention features the acquisition of a value, e.g., a composite value, that is a function of the level of gene expression of said first gene, the level of gene expression of said second gene, and a weighting factor. In an embodiment, one of said first value or said second value is a function of a weighting factor. In an embodiment, said first value is a function of a first weighting factor and said second value is a function of a second weighting factor. In an embodiment, said first weighting factor and said second weighting factor are different. In an embodiment, the invention features the acquisition of a value, e.g., a composite value, that is a function of the level of gene expression of each of a plurality of genes, and a weighting factor. In an embodiment, the invention features the acquisition of a value or values that is a function of the level of gene expression of each gene of said plurality of genes, wherein each gene in said plurality of genes is a function of a weighting factor. In an embodiment, the invention features the acquisition of a value or values that is the function of the level of gene expression of each gene of a plurality of genes that is further a function of a weighting factor. In an embodiment, the invention features the acquisition of a value or values that is the function of the level of gene expression of each gene of a plurality of genes that is further a function of a weighting factor, wherein each weighting factor for each gene of said plurality of genes is different.
In an embodiment, the invention features the acquisition of a value or values that is a function of the level of gene expression of each gene of said plurality of genes, wherein said value or values is a function of a comparison with a reference criterion. In an embodiment, said value or values is a function of the determination of whether the level of gene expression has a preselected relationship with a reference criterion, e.g., comparing said level of expression, with a preselected reference. In an embodiment, said value or values is a function of said determination; and in another embodiment, the invention features determining if said value or values has a preselected relationship with a reference criterion.
In an embodiment, the invention features the acquisition of a value or values that is a function of the level of gene expression of said plurality of genes, at a predetermined interval, e.g., a first point in time and at least a subsequent point in time.

Gene Set Score

In an embodiment, the invention features the acquisition of a gene set score. In an embodiment, the gene set score is a function of a value or values that is a function of the level of gene expression of said plurality of genes in said gene sets one and/or two and/or three and/or four and/or five and/or six and/or seven and/or eight and/or nine and/or ten and/or eleven and/or twelve and/or thirteen and/or fourteen. In an embodiment, the gene set score is a function of a value or values that is a function of the level of gene expression of said plurality of genes in said gene sets one and/or two and/or three and/or four and/or five and/or six and/or seven and/or eight and/or nine and/or ten and/or eleven and/or twelve and/or thirteen and/or fourteen and further a function of the level of gene expression of a gene or plurality of genes in Table 1.

TABLE 1

Genes of tumor initiation, EMT, and Cancer Stem Cell classifiers

DPF2	KIAA0436	CLTC	RAD51L1	STAU1	CTSL2
CASP8	CYP4V2	COPB2	EPPK1	TUBB3	CXADR
BCL2	JTV1	SLC25A25	COL1A1	UBE2S	CYP27B1
SCGN	ICMT	ECOP	MMP9	XPNPEP1	DSC2
SWAP70	DNMT3A	PDE8A	SERPINE1	CDKN1A	DSG3
KIAA0276	HNMT	STAM	SPARC	CHRD	DST
C10orf9	METTL7A	TUBB	TGFB1	H19	EPB41L4B
C10orf7	METTL2	SNX6	TGFB3	ID3	FGFBP1
ALKBH	VIL2	RAB23	TGFB1	ID4	FGFR3
TOB2	TPD52	PLAA	TGFBR1	IGFBP7	FST
XPR1	ARPC5	STC2	TGIF	LRP1	GJB3
CD59	NOL8	LTF	TGIF2	MSX1	GRHL2
LRP2	NSF	ISGF3G	THBS1	NOTCH3	HBEGF
PLP2	RAD23B	ATXN3	ANXA5	PROCR	HOOK1
MAPK14	SRP54	GTF3C3	ACTG1	GBX2	IL18
CXCL2	HSPA2	GSK3B	ARF3	KI67	IL1B
MMP7	PBP	KLF10	ATP1B3	CCNB1	IRF6
MGP	THAP2	ELL2	BAT3	BUB1	ITGB4
MLF1	CIRBP	ZBTB20	CALD1	KNTC2	JAG2
FLNB	SNRPN	IRX3	CENTD2	USP22	KLK10
SCNM1	KIAA0052	ETS1	CLIC1	HCFC1	KLK5
HSPC163	DUSP10	SERTAD1	CTBS	RNF2	KLK7
C5orf18	SSR1	MGC4251	DPYSL3	ANK3	KLK8
MGC4399	ERBB4	MAFF	DVL3	FGFR2	KRT15
CDW92	EMP1	SFPQ	EXT1	CES1	KRT16
TMC4	CHPT1	CITED4	FGFR1	COL1A2	KRT17
ZDHHC2	LRPAP1	CEBPD	FTL	COL3A1	LEPREL1
TICAM2	FLJ11752	EIF4E2	GNB2L1	COL5A2	MYO5C
KDELR3	CSTF1	HS2ST1	GPRC5A	COL6A1	NDRG1
GNPDA1	KLHL20	AGPS	H2AFZ	ANKRD25	NMU
THEM2	DNAJC13	PGK1	HIF1A	C10ORF56	PI3
DBR1	APLP2	ATIC	IL13RA1	C5ORF13	RAB25
FLJ90709	ARGBP2	ETNK1	KDELR2	KRT81	RLN2
FLJ10774	DNAJB1	LG2	LARP1	N-PAC	RNF128
C16orf33	NEBL	NCE2	LPIN2	PLEKHC1	S100A14
GAPD	SH3BGRL	8-Mar	MARS	9-Sep	S100A7
LDHA	NUDT5	CNOT4	MMP10	SYNC1	S100A8
MR-1	GABARAPL1	RNF8	MMP14	MBP	SERPINB1
LARS	MAPT	PSMA5	MT2A	ABLIM1	SERPINB2
GTPBP1	DCBLD1	DPF2	MYO10	ALDH1A3	SLC2A9
PRSS16	STK39	AMMECR1	NUP62	ALOX15B	SLPI
WFDC2	PAK2	KIAA1287	ROR1	TUBA1A	ESRP1
AIM1	CSNK2A1	LOC144233	DLC1	PPM1D	CLDN3
DHRS6	PILRB	LOC286505	GNG11	TWIST1	CLDN4
DHRS4	ERN1	PNAS-4	CDH11	FN1	ERBB3
GC15429	SGKL	FLJ20530	NR2F1	TGFBR3	SPOCK1
MGC45840	WEE1	HUMPD3	PRR16	SERPINF1	FERMT2
ECHDC2	MAST4	GC45564	MYL9	UGDH	GLYR1
GOLGIN-67	C11orf17	CAP350	DOCK10	SRGN	LTBP1
AFURS1	NUP37	ETAA16	LRIG1	FAP	FADS2
HAN11	GAS7	ZNF335	IER3	PTGER4	KANK2
DNAPTP6	TRAM2	SH3KBP1	EML1	PRKCA	PTGFR
C7orf25	BASP1	MST150	NEBL	FSTL1	COL11A2
FLJ37953	FOXO1A	PRO1073	RGL1	MMP1	KLK3
FLJ10587	POLR2A	LOC388397	MLPH	NRP1	EIF2C2
C7orf36	PER1	FKBP5	DNAJB4	FILIP1L	ZFP41
ELP4	DDIT4	HIPK2	FBLN5	SCCPDH	FAM49B
NDEL1	CD97	KLF13	RGS4	LTBP2	PSORS1C2
NPD014	BIN1	ANTXR2	HAS2	XYLT1	MRPL42
KFZP564D172	SH2B3	IFNAR1	ITGBL1	HS3ST2	MRPL54
FAM53C	DDB2	LIX1L	IGFBP4	SYT11	MRPL47
IER5	EMP3	CHST11	DPT	TSHZ1	MRPS23
LOC255783	NDST2	AKAP2	PCOLCE	THY1	EIF3S9
KIAA0146	CHST2	DTX1	GREM1	9-Sep	ALG5
KIAA0792	NT5E	ST3GAL2	PPAP2B	S100A4	DNAJC19
LOC439994	PDE4A	ADAMTS7	CDH2	TNS3	TPRXL
LOC283481	CPS1	TNRC6B	PMP22	ENOX1	NOTCH2
CG018	PTGS1	CYGB	LUM	TGFB1I1	RBM15
LOC130576	GGCX	SDHAL1	CHN1	ZEB2	ST3GAL3
NGFRAP1L1	IRF5	LOC572558	CYP1B1	LMCD1	NFYA
KIAA1217	ZBTB16	TRIO	MME	PDGFC	PCNX
4orf7	MAP4K4	FRAS1	WNT5A	ECM1	FBXO21
C21orf86	CHST7	KIAA1632	POSTN	TFPI	WWOX
C9orf64	KLF12	POLS	MMP2	TBX3	CAMK2B
FLJ13456	NFRKB	EBF	CTGF	DDR2	PNPLA2
KIAA1600	PSD	MAML2	CLIC5	PFKFB3	ANXA3
B7-H4	FKSG49	PTPRA	UGCGL1	PLOD2	AP1M2
LOC80298	NIFUN	PLEKHG2	FBXL18	PSMB7	ARTN
C7orf2	FYN	DYM	ADRBK1	PSMD8	CA2
NUCKS	ZMYM2	SOX6	SLC38A2	RIN2	CA9
DKFZP566D1346	CACNA1G	ARHGEF2	IL8RA	RYBP	CDH3
LOC388279	SLC25A16	ZCCHC6	TAS2R14	SDF4	CDS1
FLJ31795	FLII	PPP3CA	CD300LB	SETD5	COL17A1
6orf107	EIF1	FAM70B	GIPC3	SPP1	CORO1A
FLJ12439	SEPT6	TMED5	MYCBP2	LUZP1	TCHP
FLJ12806	PHF15	FLJ43663	FLJ90709	FBLN1	CDKN2C
FLJ39370	NUP188	HPS1	PCTK2	IGFBP3	VCAN
GATS	ABR	MEF2A	PDE4DIP	DCN	CD44
CCDC92	CNR1	ST3GAL5	KIAA0194	PRRX1	STARD13
FMNL2	LOC283824	SMYD3	HOM-TES-103	ANXA6	SNED1
ARID1B	FSTL4	KLF7	ENPP2	PVRL3	ZBTB38
ZFHX1B	DNM1	LOC200230	CITED2	MAP1B	SDC2
SSBP2	APOBEC3G	RERE	ZEB1	TNFAIP6	TPM1
ARID5B	ATP2B1	QKI	NID2	CYBRD1	COPZ2
LOC157381	SMPD1	BICD1	SEMA5A	FBN1	STC1
KPNA3	SLC11A1	CTNNB1	DAB2	NID1	CDH1
ARHGAP24	FXYD5	POU2F2	KCNMA1	OLFML3	KRT5
CCND2	C14orf139	EIF4ENIF1	PTX3	SNAI1	KRT6B
VIM	SH3BGRL3	BTG1	PCDH9	SNAI2	EPCAM
CREB3L1	TAGLN	CD24	BGN	SYNC	GLYR1
PALM2

Level of RNA Expression

In an embodiment, the invention features acquiring a value or values that is a function of the level of gene expression of said plurality of genes in said gene sets one and/or two and/or three and/or four and/or five and/or six and/or seven and/or eight and/or nine and/or ten and/or eleven and/or twelve and/or thirteen and/or fourteen. In an embodiment, the level of gene expression is a function of the level of RNA expression of said plurality of genes. In an embodiment, the level of gene expression is a function of the level of RNA expression of each gene of said plurality of genes. In an embodiment, the level of RNA expression is acquired. In an embodiment, the level of RNA expression of said plurality of genes is assayed. In an embodiment, the level of RNA expression is assayed by detecting an RNA product, e.g., mRNA of said sample. In an embodiment, the level of RNA expression is assayed by a hybridization based method, e.g., hybridization with a probe that is specific for said RNA product. In an embodiment, the level of RNA expression is assayed by; applying said sample, or the mRNA isolated from, or amplified from; said sample, to a nucleic acid microarray, or chip array. In an embodiment, the level of RNA expression is assayed by microarray. In an embodiment, the level of RNA expression is assayed by a polymerase chain reaction (PCR) based method, e.g., qRT-PCR. In an embodiment, the level of RNA expression is assayed by a sequencing based method. In an embodiment, the level of RNA expression is assayed by quantitative RNA sequencing. In an embodiment, the level of RNA expression is assayed by RNA in situ hybridization. In an embodiment, the level of RNA expression is assayed in the whole subject sample. In an embodiment, the level of RNA expression is assayed in a subregion of the subject sample, e.g., subregions of a tissue sample.
In an embodiment, the level of gene expression is a function of the level of protein expression of a plurality of genes in said gene sets one and/or two and/or three and/or four and/or five and/or six and/or seven and/or eight and/or nine and/or ten and/or eleven and/or twelve and/or thirteen and/or fourteen. In an embodiment, the level of gene expression is a function of the level of protein expression of said plurality of genes. In an embodiment, the level of gene expression is a function of the level of protein expression of each gene of said plurality of genes. In an embodiment, the level of protein expression is acquired. In an embodiment, the level of protein expression is assayed. In an embodiment, the level of protein expression is assayed by detecting a protein product. In an embodiment, the level of protein expression is assayed using antibodies selective for said protein product. In an embodiment, the level of protein expression is assayed by an immunohistochemistry technique. In an embodiment, the level of protein expression is assayed by an immunohistochemistry technique, using antibodies specific for said protein product. In an embodiment, the level of protein expression is assayed by an immunoassay, e.g., Western blot, enzyme linked immunosorbant assay (ELISA). In an embodiment, the level of protein expression is assayed by an immunoassay specific for said protein. In an embodiment, levels of gene expression are assessed using protein activity assays, such as functional assays. In an embodiment, the level of protein expression is assayed in the whole subject sample. In an embodiment, the level of protein expression is assayed in a subregion of the subject sample, e.g., subregions of a tissue sample.

Subject Sample

In an embodiment, the method of the invention features acquiring a subject sample, e.g., blood, urine, or tissue sample. In an embodiment, the subject sample is a tissue sample, e.g., biopsy. In an embodiment, the subject sample is a bodily fluid, e.g., blood, plasma, urine, saliva, sweat, tears, semen, or cerebrospinal fluid. In an embodiment, the subject sample is a bodily product, e.g., exhaled breath. In an embodiment, said subject sample is a tissue sample, wherein said tissue sample is derived from fixed tissue, paraffin embedded tissue, fresh tissue, or frozen tissue. In an embodiment, said subject sample is a tissue sample, wherein said tissue sample is fixed tissue, paraffin embedded tissue, fresh tissue, or frozen tissue.
In an embodiment, said subject sample is derived from a tumor. In an embodiment, said subject sample is obtained from a tumor sample. In an embodiment, said subject sample is a tumor sample. In an embodiment, said subject sample is obtained from tumor tissue. In an embodiment, the subject sample is tumor tissue. In an embodiment, said subject sample is obtained from tumor tissue, wherein said subject sample is fixed tumor tissue, paraffin embedded tumor tissue, fresh tumor tissue, or frozen tumor tissue. In an embodiment, said subject sample is a tissue sample, wherein said tissue sample is fixed, paraffin embedded, fresh, or frozen. In an embodiment, said subject sample is fixed, paraffin embedded, fresh, frozen, or fixed paraffin embedded tumor tissue.
In an embodiment, the subject sample is derived from a biopsy. In an embodiment, said subject sample derived from said biopsy is fresh tissue. In an embodiment, said subject sample derived from said biopsy is tumor tissue. In an embodiment, said subject sample derived from said biopsy is non-tumor tissue. In an embodiment, said subject sample is derived from a fine needle aspirate biopsy; large core needle biopsy; or directional vacuum assisted biopsy. In an embodiment, the subject sample is a tissue sample, wherein said tissue sample is derived from a fine needle aspirate; large core needle biopsy; or directional vacuum assisted biopsy.
In an embodiment, the subject sample is blood. In an embodiment, the subject sample is blood in which circulating tumor cells have been captured or isolated. In an embodiment, the subject sample is said circulating tumor cells that have been captured or isolated from said blood.

Location Specific Acquisition of the Level of Gene Expression

In an embodiment, the invention features, acquiring a value or values for locations in a subject sample. In an embodiment, a value or values is acquired for a plurality of locations in a subject sample. In an embodiment, a first value or values is acquired for a first location in said subject sample. In an embodiment, a second value or values is acquired for a second location in said subject sample. In an embodiment, said first value or values is different from said second value or values. In an embodiment, the invention features, determining if said first value or values and said second value or values has a preselected relationship with a reference criterion. In an embodiment, determination of whether said first value or values and/or said second value or values has a preselected relationship with a reference criterion includes comparing said first value or values with said second value or values.
In an embodiment, said first value or values is associated with an increased likelihood of comprising a cancer stem cell, cancer associated mesenchymal cell, or tumor initiating cancer cell; than is said second value or values. In an embodiment, said first value or values is associated with a higher likelihood of comprising a cancer stem cell than is said second value or values. In an embodiment, said first value or values is associated with a higher likelihood of comprising a cancer associated mesenchymal cell than is said second value or values. In an embodiment, said first value or values is associated with a higher likelihood of comprising a tumor initiating cancer cell than is said second value or values. In an embodiment, said first value or values is indicative of a cancer stem cell, cancer associated mesenchymal cell, or tumor initiating cancer cell. In an embodiment, said first value or values is indicative of a cancer stem cell. In an embodiment, said first value or values is indicative of a cancer associated mesenchymal cell. In an embodiment, said first value or values is indicative of a tumor initiating cancer cell.
In an embodiment, the invention features, classifying a location in a subject sample as a cancer stem cell, cancer associated mesenchymal cell, or tumor initiating cancer cell. In an embodiment, the invention features, classifying said location as a cancer stem cell or non-cancer stem cell. In an embodiment, the invention features, classifying said location as a cancer stem cell. In an embodiment, the invention features, classifying said location as a non-cancer stem cell. In an embodiment, the invention features, classifying said location as a cancer associated mesenchymal cell. In an embodiment, the invention features, classifying said location as a tumor initiating cancer cell. In an embodiment, the invention features, acquiring a first value or values for a first location in said subject sample, wherein responsive to said first value or values, classifying said first location as comprising a cancer stem cell or non-cancer stem cell. In an embodiment, the invention features, acquiring a first value or values for a first location in said subject sample, wherein responsive to said first value or values, classifying said first location as comprising a cancer stem cell, cancer associated mesenchymal cell, or tumor initiating cancer cell.
In an embodiment, the invention features, acquiring a first value or values for a first location in a subject sample, wherein responsive to said first value or values, classifying said first location as comprising a cancer stem cell. In an embodiment, the invention features, acquiring a first value or values for a first location in said subject sample, wherein responsive to said first value or values, classifying said first location as comprising a non-cancer stem cell. In an embodiment, the invention features, acquiring a first value or values for a first location in a subject sample, wherein responsive to said first value or values, classifying said first location as comprising a cancer associated mesenchymal cell. In an embodiment, the invention features, acquiring a first value or values for a first location in a subject sample, wherein responsive to said first value or values, classifying said first location as comprising a tumor initiating cancer cell.
In an embodiment, said first location is classified as a cancer stem cell, cancer associated mesenchymal cell, or tumor initiating cancer cell. In an embodiment, said first location is classified as a cancer stem cell. In an embodiment, said first location is classified as a cancer associated mesenchymal cell. In an embodiment, said first location is classified as a tumor initiating cancer cell. In an embodiment, said first location is classified as a non-cancer stem cell. In an embodiment, said first location comprises a cancer stem cell, cancer associated mesenchymal cell, or tumor initiating cancer cell. In an embodiment, said first location comprises a cancer stem cell. In an embodiment, said first location comprises a cancer associated mesenchymal cell. In an embodiment, said first location comprises a tumor initiating cancer cell. In an embodiment, said first location comprises a non-cancer stem cell. In an embodiment, said first location is indicative of a cancer stem cell, cancer associated mesenchymal cell, or tumor initiating cancer cell. In an embodiment, said first location is indicative of a cancer stem cell. In an embodiment, said first location is indicative of a cancer associated mesenchymal cell. In an embodiment, said first location is indicative of a tumor initiating cancer cell. In an embodiment, said first location is indicative of a non-cancer stem cell.
In an embodiment, said first location comprises a subject sample. In an embodiment, said first location comprises a whole subject sample. In an embodiment, said first location comprises a sub-region of the subject sample. In an embodiment, said first location and said second location are separated by zero microns, i.e., said first location and second location are adjoining. In an embodiment, said first location and said second location are separated by more than zero microns; by more than ten microns; by more than twenty microns; by more than thirty microns; by more than forty microns; by more than fifty microns; by more than sixty microns; by more than seventy microns; by more than eighty microns; by more than ninety microns; or by more than one hundred microns. In an embodiment, said first location and said second location are separated by more than one thousand microns. In an embodiment, said first location and said second location are separated by at least ten microns; in an embodiment, said first location and said second location are separated by at least twenty microns; by at least thirty microns; by at least forty microns; by at least fifty microns; by at least sixty microns; by at least seventy microns; by at least eighty microns; by at least ninety microns; or by at least one hundred microns. In an embodiment, said first location and said second location are separated by more than one hundred microns. In an embodiment, said first location and said second location are separated by more than two hundred microns; three hundred microns; four hundred microns; five hundred microns; six hundred microns; seven hundred microns; eight hundred microns; nine hundred microns; or one thousand microns. In an embodiment, said first location and said second location are separated by at least one thousand microns. In an embodiment, said first location and said second location are separated by the maximum distance two locations of said subject sample can be separated. In an embodiment, said first location and said second location are separated by a distance between and including, zero and the maximum distance two locations of said subject sample can be separated.
In an embodiment, the average distance between said first location and said second location is more than zero microns; in an embodiment, the average distance between said first location and said second location is approximately ten microns; approximately twenty microns; approximately thirty microns; approximately forty micron; approximately fifty microns; approximately sixty microns; approximately seventy microns; approximately eighty microns; approximately ninety microns; or approximately one hundred microns. In an embodiment, the average distance between said first location and said second location is more than approximately fifty microns.
In an embodiment, the average distance between said first location and said second location is zero microns; in an embodiment, the average distance between said first location and said second location is more than ten microns; more than twenty microns; more than thirty microns; more than forty micron; more than fifty microns; more than sixty microns; more than seventy microns; more than eighty microns; more than ninety microns; or more than one hundred microns.
In an embodiment, the average distance between said first location and said second location is more than approximately one hundred microns. In an embodiment, the average distance between said first location and said second location is more than approximately two hundred; more than approximately three hundred; more than approximately four hundred; more than approximately five hundred; more than approximately six hundred; more than approximately seven hundred; more than approximately eight hundred; more than approximately nine hundred; or more than approximately one thousand microns. In an embodiment, the average distance between said first location and said second location is more than one thousand microns.
In an embodiment, the average distance between said first location and said second location is at least approximately ten microns; at least approximately twenty microns; at least approximately thirty microns; at least approximately forty microns; at least approximately fifty microns; at least approximately sixty microns; at least approximately seventy microns; at least approximately eighty microns; at least approximately ninety microns; at least approximately one hundred microns; at least approximately two hundred microns.
In an embodiment, said first value or values of said first location is a function of the level of gene expression of a gene at said first location. In embodiment, said first value or values is a function of the level of gene expression of a plurality of genes at said first location. In an embodiment, said first value or values is a function of the level of gene expression of each gene isoform of a plurality of genes at said first location. In an embodiment, the invention features the first value or values of said first location is a function of the level of gene expression of a gene or a plurality of genes at said first location, and responsive to said first value or values classifying said first location as a cancer stem cell or non cancer stem cell. In an embodiment, the invention features the first value or values of said first location is a function of the level of gene expression of a gene or a plurality of genes at said first location, and responsive to said first value or values classifying said first location as a cancer stem cell, cancer associated mesenchymal cell, or tumor initiating cancer cell. In an embodiment, said gene or said plurality of genes is in Table 1. In an embodiment, the level of gene expression is a function of the level of RNA expression of said gene or said plurality of genes. In an embodiment, the level of RNA expression of said gene or plurality of genes is assayed. In an embodiment, the level of RNA expression is assayed by detecting an RNA product. In an embodiment, the level of RNA expression is assayed by RNA in situ hybridization. In an embodiment, the level of gene expression is a function of the level of protein expression of said gene or said plurality of genes. In an embodiment, the level of protein expression is acquired. In an embodiment, the level of protein expression is assayed. In an embodiment, the level of protein expression is assayed by detecting a protein product. In an embodiment, the level of protein expression is assayed using antibodies selective for said protein product. In an embodiment, the level of protein expression is assayed by immunohistochemistry.

Administration

In an embodiment, the invention features administering an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells. In an embodiment, said agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells or cancer stem cells is administered to said subject. In an embodiment, the agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells is selected from, e.g., salinomycin; a gamma secretase inhibitor; a DLL4 inhibitor, e.g., a therapeutic antibody targeting DLL4; a TRAIL inhibitor, e.g., a therapeutic antibody targeting TRAIL; a Hedgehog inhibitor, e.g., a therapeutic antibody targeting Hedgehog; a NOTCH3 inhibitor, e.g., a therapeutic antibody targeting NOTCH3; a NOTCH4 inhibitor, e.g., a therapeutic antibody targeting NOTCH4; a panNOTCH inhibitor, e.g., a therapeutic antibody targeting panNOTCH; a FGFR1 inhibitor, e.g., a therapeutic antibody targeting FGR1; a FGFR2 inhibitor, e.g., a therapeutic antibody targeting FGR2; a FGFR3 inhibitor, e.g., a therapeutic antibody targeting FGR3; a FGFR4 inhibitor, e.g., a therapeutic antibody targeting FGR4; a RON inhibitor, e.g., a therapeutic antibody targeting RON; Wnt pathway inhibitor, e.g., therapeutic antibodies targeting the Wnt pathway; a PI3Kinase inhibitor; a mTOR inhibitor; sodium meta arsenite; verapail; reserpine; a perifosen inhibitor of FAK1; a FAK inhibitor; a p38 inhibitor.
In an embodiment, the method features selecting a regimen, e.g., dosage, formulation, route of administration, number of dosages, or adjunctive therapies, of the agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells. In an embodiment, said selecting is responsive to said value or values that is a function of the level of gene expression of said plurality of genes selected from said first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or seven and/or eighth gene sets. In an embodiment, the invention features administering an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells to the subject according to the selected regimen. In an embodiment, said administration is provided responsive to acquiring knowledge or information of said value or values from another party. In an embodiment, said administration is provided responsive to an identification of said value or values, wherein said identification arises from collaboration with another party. In an embodiment, the invention features receiving a communication of the presence of said value or values that is a function of the level of gene expression for each of a plurality of genes selected from said first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or seventh and/or eighth and/or ninth and/or tenth and/or eleventh and/or twelfth and/or thirteenth and/or fourteenth set of genes in a subject. In an embodiment, the acquisition of said value or values is at the time of or after diagnosis of cancer in said subject. In an embodiment, the acquisition of said value or values is post diagnosis of said cancer in the subject. In an embodiment, said subject has cancer. In an embodiment, the cancer is characterized as comprising cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells. In an embodiment, the cancer is characterized as comprising cancer associated mesenchymal cells. In an embodiment, the cancer is characterized as comprising tumor initiating cancer cells. In an embodiment, the cancer is characterized as comprising cancer stem cells. In an embodiment, the cancer is characterized as being enriched with cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells. In an embodiment, the cancer is characterized as being enriched with cancer associated mesenchymal cells. In an embodiment, the cancer is characterized as being enriched with tumor initiating cancer cells. In an embodiment, the cancer is characterized as being enriched with cancer stem cells.
In an embodiment, said cancer is an epithelial cell cancer. In an embodiment, said cancer is breast, lung, pancreatic, colorectal, prostate, head and neck, melanoma, acute myelogenous leukemia, glioblastoma, triple negative breast cancer, basal-like breast cancer, or claudin-low breast cancer. In another embodiment, said cancer is breast cancer. In an embodiment, said cancer is triple negative breast cancer. In an embodiment, the cancer is basal-like breast cancer. In an embodiment, the cancer is claudin-low breast cancer. In an embodiment, said cancer is recurrent, i.e., cancer that returns following treatment, and after a period of time in which said cancer was undetectable. In another embodiment, said cancer is a primary tumor, i.e., located at the anatomical site of tumor growth initiation. In an embodiment, said cancer is metastatic, i.e., appearing at a second anatomical site other than the anatomical site of tumor growth initiation.
In an embodiment of the invention, the value or values that is a function of the level of gene expression of said plurality of genes selected from said first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or seventh and/or eighth and/or ninth and/or tenth and/or eleventh and/or twelfth and/or thirteenth and/or fourteenth set of genes; is acquired prior to, during, or after administration of a treatment to said subject. In an embodiment, said value or values is acquired prior to the administration of a treatment to said subject. In an embodiment, said value or values is acquired during the administration of a treatment to said subject. In an embodiment, said value or values is acquired after the administration of a treatment to said subject. In an embodiment, said subject is a non-responder, to said treatment. In an embodiment, said treatment is an anti-cancer treatment, e.g., chemotherapeutic agent, radiation treatment, surgery, etc. In an embodiment, said anti-cancer treatment is a chemotherapeutic agent. In an embodiment, said chemotherapeutic agent may include but is not limited to is one or more of the following chemotherapeutic agents: alkylating agents (e.g., nitrogen mustards such as chlorambucil, cyclophosphamide, isofamide, mechlorethamine, melphalan, and uracil mustard; aziridines such as thiotepa; methanesulphonate esters such as busulfan; nitroso ureas such as carmustine, lomustine, and streptozocin; platinum complexes such as cisplatin and carboplatin; bioreductive alkylators such as mitomycin, procarbazine, dacarbazine and altretamine); DNA strand-breakage agents (e.g., bleomycin); topoisomerase II inhibitors (e.g., amsacrine, dactinomycin, daunorubicin, idarubicin, mitoxantrone, doxorubicin, etoposide, and teniposide); DNA minor groove binding agents (e.g., plicamydin); antimetabolites (e.g., folate antagonists such as methotrexate and trimetrexate; pyrimidine antagonists such as fluorouracil, fluorodeoxyuridine, CB3717, azacitidine, cytarabine, and floxuridine; purine antagonists such as mercaptopurine, 6-thioguanine, fludarabine, pentostatin; asparginase; and ribonucleotide reductase inhibitors such as hydroxyurea); tubulin interactive agents (e.g., vincristine, vinblastine, and paclitaxel (Taxol)); hormonal agents (e.g., estrogens; conjugated estrogens; ethinyl estradiol; diethylstilbesterol; chlortrianisen; idenestrol; progestins such as hydroxyprogesterone caproate, medroxyprogesterone, and megestrol; and androgens such as testosterone, testosterone propionate, fluoxymesterone, and methyltestosterone); adrenal corticosteroids (e.g., prednisone, dexamethasone, methylprednisolone, and prednisolone); leutinizing hormone releasing agents or gonadotropin-releasing hormone antagonists (e.g., leuprolide acetate and goserelin acetate); and antihormonal antigens (e.g., tamoxifen, antiandrogen agents such as flutamide; and antiadrenal agents such as mitotane and aminoglutethimide). In an embodiment, said chemotherapeutic agent is selected from one or more of the following chemotherapeutic agents: Capecitabine, Carboplatin, Cisplatin, Cyclophosphamide, Docetaxel, Doxorubicin, Epirubicin, Eribulin, mesylate5-Fluorouracil, Gemcitabine, Ixabepilone, Liposomal doxorubicin, Methotrexate, Paclitaxel, or Vinorelbine; or any combination thereof.
In an embodiment, the invention features administering an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells and a second treatment. In an embodiment, said second treatment is an anti-cancer agent. In an embodiment, said second treatment is an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells. In an embodiment, said second treatment is not an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells. In an embodiment, said second treatment kills or inhibits growth of non-cancer stem cells in the subject. In an embodiment, the second treatment kills or inhibits growth of cancer cells that are not cancer stem cells, cancer associated mesenchymal cells, or tumor initiating cancer cells. In an embodiment, the second treatment is an anti-cancer treatment that does not target cancer stem cells, cancer associated mesenchymal cells, or cancer stem cells. In an embodiment, the second treatment is an anti-cancer treatment that does not primarily target cancer stem cells, cancer associated mesenchymal cells, or cancer stem cells. In an embodiment, said second treatment kills or inhibits growth of non-cancer associated mesenchymal cells, non-tumor initiating cancer cells, or non-cancer stem cells in the subject. In an embodiment, said second treatment is a chemotherapeutic agent. In an embodiment, said second treatment may include but is not limited to one or more of the following: alkylating agents (e.g., nitrogen mustards such as chlorambucil, cyclophosphamide, isofamide, mechlorethamine, melphalan, and uracil mustard; aziridines such as thiotepa; methanesulphonate esters such as busulfan; nitroso ureas such as carmustine, lomustine, and streptozocin; platinum complexes such as cisplatin and carboplatin; bioreductive alkylators such as mitomycin, procarbazine, dacarbazine and altretamine); DNA strand-breakage agents (e.g., bleomycin); topoisomerase II inhibitors (e.g., amsacrine, dactinomycin, daunorubicin, idarubicin, mitoxantrone, doxorubicin, etoposide, and teniposide); DNA minor groove binding agents (e.g., plicamydin); antimetabolites (e.g., folate antagonists such as methotrexate and trimetrexate; pyrimidine antagonists such as fluorouracil, fluorodeoxyuridine, CB3717, azacitidine, cytarabine, and floxuridine; purine antagonists such as mercaptopurine, 6-thioguanine, fludarabine, pentostatin; asparginase; and ribonucleotide reductase inhibitors such as hydroxyurea); tubulin interactive agents (e.g., vincristine, vinblastine, and paclitaxel (Taxol)); hormonal agents (e.g., estrogens; conjugated estrogens; ethinyl estradiol; diethylstilbesterol; chlortrianisen; idenestrol; progestins such as hydroxyprogesterone caproate, medroxyprogesterone, and megestrol; and androgens such as testosterone, testosterone propionate, fluoxymesterone, and methyltestosterone); adrenal corticosteroids (e.g., prednisone, dexamethasone, methylprednisolone, and prednisolone); leutinizing hormone releasing agents or gonadotropin-releasing hormone antagonists (e.g., leuprolide acetate and goserelin acetate); and antihormonal antigens (e.g., tamoxifen, antiandrogen agents such as flutamide; and antiadrenal agents such as mitotane and aminoglutethimide). In an embodiment, said second therapeutic agent is selected from Capecitabine, Carboplatin, Cisplatin, Cyclophosphamide, Docetaxel, Doxorubicin, Epirubicin, Eribulin, mesylate5-Fluorouracil, Gemcitabine, Ixabepilone, Liposomal doxorubicin, Methotrexate, Paclitaxel, or Vinorelbine; or any combination thereof. In an embodiment, the invention features further administering an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cells, or cancer stem cells and more than one additional therapeutic agent.
In an embodiment, the invention includes, responsive to the acquisition of said value or values that is a function of the level of gene expression of said plurality of genes selected from said first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or seventh and/or eighth and/or ninth and/or tenth and/or eleventh and/or twelfth and/or thirteenth and/or fourteenth set of genes; further stratifying a patient population. In an embodiment, the invention features, responsive to the acquisition of said value or values; further identifying or selecting said subject as likely or unlikely to respond positively to a treatment. In another embodiment, the invention features, responsive to the acquisition of said value or values; further selecting a treatment. In another embodiment, the invention features, responsive to the acquisition of said value or values; further prognosticating the time course of the disease in the subject. In an embodiment, said disease is a cancer. In an embodiment, the invention features, responsive to the acquisition of said value or values, one or more of the following: stratifying a patient population; identifying or selecting said subject as likely or unlikely to respond to a treatment; selecting a treatment option; prognosticating the time course of the disease in the subject; measuring the response at the end of therapy and predicting the long term outcome; and/or determining the cancer stem cell population as a predictor of response to a treatment or therapy.

Genotype

In an embodiment, the method of the invention features the acquisition of a genotype of said subject sample. The subject sample can be any suitable subject sample including those subject samples previously mentioned. In an embodiment, said subject sample is a tumor sample. In an embodiment, at least one nucleotide of the subject sample is sequenced to determine the presence or absence of at least one genetic event associated with cancer. In an embodiment, at least one oncogene or tumor suppressor gene in the sample is sequenced. In an embodiment, the oncogene or oncogenes or tumor suppressor gene or tumor suppressor genes may include but is not limited to one or any combination of: Abl, Af4/hrx, akt-2, alk, alk/npm, aml 1, aml 1/mtg8, APC, axl, bcl-2, bcl-3, bcl-6, bcr/abl, brca-1, brca-2, beta-catenin, CDKN2, c-myc, c-sis, dbl, dek/can, E2A/pbx1, egfr, enl/hrx, erg/TLS, erbB, erbB-2, erk, ets-1, ews/fli-1, fms, fos, fps, gli, gsp, HER2/neu, hox11, hst, IL-3, int-2, jun, kit, KS3, K-sam, Lbc, lck, lmo1, lmo2, L-myc, li1-1, lyt-10, lyt-10/C alpha1, mas, mdm-2, mll, mos, mtg8/aml1, myb, myc, MYH11/CBFB, neu, nm23, N-myc, ost, p53, pax-5, pbx1/E2A, pdgfr, PI3-K, pim-1, PRAD-1, raf, RAR/PML, rash, rasK, rasN, Rb, rel/nrg, ret, rhom1, rhom2, ros, ski, sis, set/can, src, tal1, tal2, tan-1, telomerase, Tiam1, TSC2, trk, vegfr, or wnt.

Reports

In an embodiment, the present invention features optionally providing a prediction of the likelihood that a subject will respond positively or will not respond positively to treatment with an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells. In an embodiment, said prediction is in the form of a report. In an embodiment, said predication includes a recommendation of whether said subject should be treated with a preselected drug, or treatment with a preselected drug should be withheld. In an embodiment, said preselected drug is an anti-cancer agent. In an embodiment, said preselected drug is an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells. In an embodiment, said agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells is selected from: e.g., salinomycin; a gamma secretase inhibitor; a DLL4 inhibitor, e.g., a therapeutic antibody targeting DLL4; a TRAIL inhibitor, e.g., a therapeutic antibody targeting TRAIL; a Hedgehog inhibitor, e.g., a therapeutic antibody targeting Hedgehog; a NOTCH3 inhibitor, e.g., a therapeutic antibody targeting NOTCH3; a NOTCH4 inhibitor, e.g., a therapeutic antibody targeting NOTCH4; a panNOTCH inhibitor, e.g., a therapeutic antibody targeting panNOTCH; a FGFR1 inhibitor, e.g., a therapeutic antibody targeting FGR1; a FGFR2 inhibitor, e.g., a therapeutic antibody targeting FGR2; a FGFR3 inhibitor, e.g., a therapeutic antibody targeting FGR3; a FGFR4 inhibitor, e.g., a therapeutic antibody targeting FGR4; a RON inhibitor, e.g., a therapeutic antibody targeting RON; Wnt pathway inhibitor, e.g., therapeutic antibodies targeting the Wnt pathway; a PI3Kinase inhibitor; a mTOR inhibitor; sodium meta arsenite; verapail; reserpine; a perifosen inhibitor of FAK1; a FAK inhibitor; a p38 inhibitor.

Kits or Products

In an aspect, the present invention includes a kit or product comprising a first agent capable of interacting with a gene expression product of a gene from a first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or seventh and/or eighth and/or ninth and/or tenth and/or eleventh and/or twelfth and/or thirteenth and/or fourteenth set of genes.
In an embodiment, the first set of genes are related to the delineation of a cancer stem cell, cancer associated mesenchymal cell, or a tumor initiating cancer cell phenotype and the determination of the attributes of the EMT in cancers. For example,
(i) said first set of genes comprises or consists of:

- KLF13, ANTXR2, IFNAR1, LIX1L, CHST11, AKAP2, PALM2-AKAPT, LOC80298, C7orf2, NUCKS, DKFZP566D1346, LOC388279, FLJ31795, 6orf107, FLJ12439, ENPP2, CITED2, ZEB1, NID2, SEMA5A, DAB2, KCNMA1, PTX3, PCDH9, CREB3L1, BGN, ROR1, DLC1, GNG11, CDH11, NR2F1, PRR16, MYL9, DOCK10, LRIG1, IER3, EML1, NEBL, RGL1, MLPH, DNAJB4, FBLN5, RGS4, HAS2, ITGBL1, IGFBP4, DPT, PCOLCE, GREM1, PPAP2B, CDH2, PMP22, LUM, CHN1, CYP1B1, MME, WNT5A, POSTN, MMP2, CTGF, FBLN1, IGFBP3, DCN, PRRX1, ANXA6, PVRL3, MAP1B, TNFAIP6, CYBRD1, FBN1, NID1, OLFML3, SNAI1, SNAI2, TAGLN, SYNC, TUBA1A, PPM1D, TWIST1, FN1, TGFBR3, SERPINF1, UGDH, SRGN, FAP, PTGER4, PRKCA, FSTL1, MMP1, NRP1, FILIP1L, SCCPDH, LTBP2, XYLT1, HS3ST2, SYT11, TSHZ1, THY1, Sep9, S100A4, TNS3, ENOX1, TGFB1I1, ZEB2, LMCD1, PDGFC, ECM1, TFPI, TBX3, DDR2, PTGER2, CDKN2C, VCAN, CD44, STARD13, SNED1, ZBTB38, SDC2, TPM1, COPZ2, STC1, CDH1, KRT5, KRT6B, CD24, EPCAM, ESRP1, CLDN3, CLDN4, ERBB3, SPOCK1, FERMT2, GLYR1, LTBP1, FADS2, KANK2, PTGFR, COL11A2, KLK3, EIF2C2, ZFP41, FAM49B, PSORS1C2, MRPL42, MRPL54, MRPL47, MRPS23, EIF3S9, ALG5, DNAJC19, TPRXL, NOTCH2, RBM15, ST3GAL3, NFYA, PCNX, FBXO21, WWOX, CAMK2B, PNPLA2, CLIC5, UGCGL1, FBXL18, ADRBK1, SLC38A2, IL8RA, TAS2R14, CD300LB, GIPC3, RAD51L1, EPPK1, COL1A1, MMP9, SERPINE1, SPARC, TGFB1, TGFB3, TGFBI, TGFBR1, TGIF, TGIF2, THBS1, ANXA5, ACTG1, ARF3, ATP1B3, BAT3, CALD1, CENTD2, CLIC1, CTBS, DPYSL3, DVL3, EXT1, FGFR1, FTL, GNB2L1, GPRC5A, H2AFZ, HIF1A, IL13RA1, KDELR2, LARP1, LPIN2, MARS, MMP10, MMP14, MT2A, MYO10, NUP62, PFKFB3, PLOD2, PSMB7, PSMD8, RIN2, RYBP, SDF4, SETD5, SPP1, STAU1, TUBB3, UBE2S, XPNPEP1, CDKN1A, CHRD, H19, ID3, ID4, IGFBP7, LRP1, MSX1, NOTCH3, PROCR, GBX2, KI67, CCNB1, BUB1, KNTC2, USP22, HCFC1, RNF2, ANK3, FGFR2, CES1, COL1A2, COL3A1, COL5A2, COL6A1, ANKRD25, C10ORF56, C5ORF13, KRT81, NPAC, PLEKHC1, SEPT9, SYNC1, MBP, ABLIM1, ALDH1A3, ALOX15B, ANXA3, AP1M2, ARTN, CA2, CA9, CDH3, CDS1, COL17A1, CORO1A, CTSL2, CXADR, CYP27B1, DSC2, DSG3, DST, EPB41L4B, FGFBP1, FGFR3, FST, GJB3, GRHL2, HBEGF, HOOK1, IL18, IL1B, IRF6, ITGB4, JAG2, KLK10, KLK5, KLK7, KLK8, KRT15, KRT16, KRT17, LEPREL1, MYO5C, NDRG1, NMU, PI3, RAB25, RLN2, RNF128, S100A14, S100A7, S100A8, SERPINB1, SERPINB2, SLC2A9, SLPI, SNX10, SPINT2, SPRR1A, SPRR1B, ST14, SYK, TMEM40, TRIM29, TSPAN1, UCHL1, VSNL1, ZBED2, AQP3, CBARA1, CLCA2, CNTNAP2, CSTA, DSC3, ESRP2, FXYD3, GJB5, GPR87, IL1A, LAMA3, LAMB3, MAP7, MAPK13, MPZL2, NRG1, SCNN1A, SERPINB13, SLC7A5, SPINT1, ST6GALNAC2, TGFA, TP63, TPD52L1, VGLL1, SLC6A8, ABCA12, ANXA8, ANXA8L1, ARHGAP25, ARHGAP8, BDKRB2, BIK, C10ORF10, C10ORF116, C1ORF116, C20ORF19, C6ORF105, CELSR2, CKMT1A, CKMT1B, CLDN1, CST6, CYP4F11, E2F5, ELMO3, EPHA1, EVA1, EXPH5, F11R, FAT2, FLJ12684, FLJ20366, FZD3, GLS2, GNAL, IFI30, IGFBP2, IL1RN, IL4R, IRX4, JUP, KCNK1, KIAA0040, KIAA0888, KIAA1815, KRT14, KRT18, LAD1, LGALS7, LOC553158, LOC653562, LOC728113, LOC729884, LRRC1, LSR, MST1R, MYO1D, NAIP, OCLN, NEFM, NUP62CL, PERP, PLS1, POLR3G, PRKCH, PRRG4, PRSS8, PTPN3, RBM35A, RBM35B, RHBDF2, RHOD, RPS6KA1, RTEL1, SAA1, SAA1, SAA2, SH2D3A, SLC6A10P, SMPDL3B, SNCA, STAC, STAP2, TACSTD1, THBD, TMEM30B, TMPRSS11E, TMPRSS4, TNFRSF6B, TP73L, WWC1, XDH, ZNF165, AXL, GPR30, PAX2, PI3KA, CD133, ALDH1, BMI1, KRT19, MSI1, JAG1, p38, ESA1, FOXC2, FOXC1, TWIST2, GSC, TCF3, MLPH1, ARHGEF11a, DPF2, CASP8, BCL2, SCGN, SWAP70, KIAA0276, C10orf9, C10orf7, ALKBH, TOB2, XPR1, CD59, LRP2, PLP2, MAPK14, CXCL2, MMP7, MGP, MLF1, FLNB, SCNM1, HSPC163, C5orf18, MGC4399, CDW92, TMC4, ZDHHC2, TICAM2, KDELR3, GNPDA1, THEM2, DBR1, FLJ90709, FLJ10774, C16orf33, GAPD, LDHA, MR-1, LARS, GTPBP1, PRSS16, WFDC2, AIM1, DHRS6, DHRS4, GC15429, MGC45840, ECHDC2, GOLGIN-67, AFURS1, KIAA0436, CYP4V2, JTV1, ICMT, DNMT3A, HNMT, METTL7A, METTL2, VIL2, TPD52, ARPC5, NOL8, NSF, RAD23B, SRP54, HSPA2, PBP, THAP2, CIRBP, SNRPN, KIAA0052, DUSP10, SSR1, ERBB4, EMP1, CHPT1, LRPAP1, FLJ11752, CSTF1, KLHL20, DNAJC13, APLP2, ARGBP2, DNAJB1, NEBL, SH3BGRL, NUDT5, GABARAPL1, MAPT, DCBLD1, STK39, PAK2, CSNK2A1, PILRB, ERN1, SGKL, WEE1, MAST4, C11orf17, NUP37, CLTC, COPB2, SLC25A25, ECOP, PDE8A, STAM, TUBB, SNX6, RAB23, PLAA, STC2, LTF, ISGF3G, ATXN3, GTF3C3, GSK3B, KLF10, ELL2, ZBTB20, IRX3, ETS1, SERTAD1, MGC4251, MAFF, SFPQ, CITED4, CEBPD, EIF4E2, HS2ST1, AGPS, PGK1, ATIC, ETNK1, LG2, NCE2, MARCH8, CNOT4, RNF8, PSMA5, DPF2, AMMECR1, KIAA1287, LOC144233, LOC286505, PNAS-4, FLJ20530, HUMPD3, GC45564, CAP350, ETAA16, HAN11, DNAPTP6, C7orf25, FLJ37953, FLJ10587, C7orf36, ELP4, NDEL1, NPD014, KFZP564D172, FAM53C, IER5, LOC255783, KIAA0146, KIAA0792, LOC439994, LOC283481, CG018, LOC130576, NGFRAP1L1, KIAA1217, 4orf7, C21orf86, C9orf64, FLJ13456, FLJ12806, FLJ39370, GATS, CCDC92, FMNL2, ARID1B, ZFHX1B, SSBP2, ARID5B, LOC157381, KPNA3, ARHGAP24, CCND2, VIM, MYCBP2, GAS7, TRAM2, BASP1, FOXO1A, POLR2A, PER1, DDIT4, CD97, BIN1, SH2B3, DDB2, EMP3, NDST2, CHST2, NT5E, PDE4A, CPS1, PTGS1, GGCX, IRF5, ZBTB16, MAP4K4, CHST7, KLF12, NFRKB, PSD, FKSG49, NIFUN, FYN, ZMYM2, CACNA1G, SLC25A16, FLII, EIF1, SEPT6, NPAC, PHF15, NUP188, ABR, CNR1, LOC283824, FSTL4, DNM1, APOBEC3G, ATP2B1, SMPD1, SLC11A1, FXYD5, C14orf139, SH3BGRL3, LUZP1, ZNF335, SH3 KBP1, MST150, PRO1073, LOC388397, FKBP5, HIPK2, DTX1, ST3GAL2, ADAMTS7, TNRC6B, CYGB, SDHAL1, LOC572558, TRIO, FRAS1, KIAA1632, POLS, EBF, MAML2, PTPRA, PLEKHG2, DYM, SOX6, ARHGEF2, ZCCHC6, PPP3CA, FAM70B, TMED5, FLJ43663, HPS1, MEF2A, ST3GAL5, SMYD3, KLF7, LOC200230, RERE, QKI, BICD1, CTNNB1, POU2F2, EIF4ENIF1, BTG1, TCHP, FLJ90709, PCTK2, KIAA1600, B7-H4, PDE4DIP, KIAA0194, HOM-TES; and
- (a) BGN, BIN1, CDH11, CDH2, CHN1, COL1A2, COL3A1, COL5A2, CREB3L1, CTGF, CYBRD1, DCN, DLC1, DNAJB4, DPT, ENPP2, FBLN1, FBLN5, FBN1, GREM1, HAS2, IGFBP3, IGFBP4, MLPH, MME, MMP2, MYL9, NEBL, NID1, NR2F1, OLFML3, PCOLCE, PMP22, POSTN, PPAP2B, PRR16, PRRX1, PTX3, PVRL3, RGL1, RGS4, ROR1, SEMA5A, TNFAIP6, TRAM2, WNT5A, ZEB1, ANXA6, CITED2, COL1A1, COL6A1, CYP1B1, DAB2, DOCK10, EML1, GNG11, IER3, ITGBL1, KCNMA1, LRIG1, LUM, MAP1B, NID2, PCDH9, PRKCA, PTGER4, PTGFR, SDC2, SERPINF1, SNED1, SYT11, TFPI, TGFBR3, VCAN, ANKRD25, C10ORF56, C5ORF13, CCDC92, CDKN2C, COPZ2, DDR2, ECM1, ENOX1, FADS2, FAP, FILIP1L, FSTL1, HS3ST2, KRT81, LMCD1, LTBP1, LTBP2, MMP1, NPAC, NRP1, PDGFC, PLEKHC1, PPM1D, PTGER2, SCCPDH, SEPT9, SPOCK1, SRGN, STARD13, STC1, SYNC1, TAGLN, TBX3, TGFB1I1, THY1, TNS3, TPM1, TSHZ1, TUBA1A, UGDH, VIM, XYLT1, ZBTB38, MBP, ABLIM1, ALDH1A3, ALOX15B, ANXA3, AP1M2, ARTN, CA2, CA9, CAMK2B, CCND2, CDH1, CDH3, CDS1, COL17A1, CORO1A, CTSL2, CXADR, CYP27B1, DSC2, DSG3, DST, EPB41L4B, FGFBP1, FGFR3, FST, GJB3, GRHL2, HBEGF, HOOK1, IL18, IL1B, IRF6, ITGB4, JAG2, KLK10, KLK5, KLK7, KLK8, KRT15, KRT16, KRT17, KRT5, KRT6B, LEPREL1, MYO5C, NDRG1, NMU, PI3, RAB25, RLN2, RNF128, S100A14, S100A7, S100A8, SERPINB1, SERPINB2, SLC2A9, SLPI, SNX10, SPINT2, SPRR1A, SPRR1B, ST14, SYK, TMEM40, TRIM29, TSPAN1, UCHL1, VSNL1, ZBED2, AQP3, CBARA1, CD24, CLCA2, CNTNAP2, CSTA, DSC3, EPCAM, ESRP2, FXYD3, GJB5, GPR87, IL1A, LAMA3, LAMB3, MAP7, MAPK13, MMP9, MPZL2, NRG1, SCNN1A, SERPINB13, SLC7A5, SPINT1, ST6GALNAC2, TGFA, TP63, TPD52L1, VGLL1, SLC6A8, ABCA12, ANK3, ANXA8, ANXA8L1, ARHGAP25, ARHGAP8, BDKRB2, BIK, C10ORF10, C10ORF116, C1ORF116, C20ORF19, C6ORF105, CELSR2, CKMT1A, CKMT1B, CLDN1, CST6, CYP4F11, E2F5, ELMO3, EPHA1, EVA1, EXPH5, F11R, FAT2, FGFR2, FLJ12684, FLJ20366, FZD3, GLS2, GNAL, IFI30, IGFBP2, IL1RN, IL4R, IRX4, JUP, KCNK1, KIAA0040, KIAA0888, KIAA1815, KRT14, KRT18, LAD1, LGALS7, LOC553158, LOC653562, LOC728113, LOC729884, LRRC1, LSR, MST1R, MYO1D, NAIP, OCLN, NEFM, NUP62CL, PERP, PLS1, POLR3G, PRKCH, PRRG4, PRSS8, PTPN3, RBM35A, RBM35B, RHBDF2, RHOD, RPS6KA1, RTEL1, SAA1, SAA1, SAA2, SH2D3A, SLC6A10P, SMPDL3B, SNCA, STAC, STAP2, TACSTD1, THBD, TMEM30B, TMPRSS11E, TMPRSS4, TNFRSF6B, TP73L, WWC1, XDH, ZNF165; and
- (b) BGN, BIN1, CDH11, CDH2, CHN1, COL1A2, COL3A1, COL5A2, CREB3L1, CTGF, CYBRD1, DCN, DLC1, DNAJB4, DPT, ENPP2, FBLN1, FBLN5, FBN1, GREM1, HAS2, IGFBP3, IGFBP4, MLPH, MME, MMP2, MYL9, NEBL, NID1, NR2F1, OLFML3, PCOLCE, PMP22, POSTN, PPAP2B, PRR16, PRRX1, PTX3, PVRL3, RGL1, RGS4, ROR1, SEMA5A, TNFAIP6, TRAM2, WNT5A, ZEB1, ANXA6, CITED2, COL1A1, COL6A1, CYP1B1, DAB2, DOCK10, EML1, GNG11, IER3, ITGBL1, KCNMA1, LRIG1, LUM, MAP1B, NID2, PCDH9, PRKCA, PTGER4, PTGFR, SDC2, SERPINF1, SNED1, SYT11, TFPI, TGFBR3, VCAN, ANKRD25, C10ORF56, C5ORF13, CCDC92, CDKN2C, COPZ2, DDR2, ECM1, ENOX1, FADS2, FAP, FILIP1L, FSTL1, HS3ST2, KRT81, LMCD1, LTBP1, LTBP2, MMP1, NPAC, NRP1, PDGFC, PLEKHC1, PPM1D, PTGER2, SCCPDH, SEPT9, SPOCK1, SRGN, STARD13, STC1, SYNC1, TAGLN, TBX3, TGFB1I1, THY1, TNS3, TPM1, TSHZ1, TUBA1A, UGDH, VIM, XYLT1, ZBTB38; and
- (c) ENPP2, CITED2, ZEB1, NID2, SEMA5A, DAB2, KCNMA1, PTX3, PCDH9, CREB3L1, BGN, ROR1, DLC1, GNG11, CDH11, NR2F1, NR2F2, PRR16, MYL9, DOCK10, LRIG1, IER3, EML1, NEBL, RGL1, MLPH, TRAM2, DNAJB4, FBLN5, RGS4, HAS2, ITGBL1, IGFBP4, BIN1, DPT, PCOLCE, GREM1, PPAP2B, CDH2, PMP22, LUM, CHN1, CYP1B1, MME, WNT5A, POSTN, MMP2, CTGF, FBLN1, IGFBP3, DCN, PRRX1, ANXA6, PVRL3, MAP1B, TNFAIP6, CYBRD1, FBN1, NID1, OLFML3, SNAI1, SNAI2, TAGLN, SYNC, TUBA1A, PPM1D, TWIST1, FN1, TGFBR3, SERPINF1, UGDH, SRGN, FAP, PTGER4, VIM, PRKCA, FSTL1, MMP1, NRP1, FILIP1L, SCCPDH, LTBP2, XYLT1, HS3ST2, SYT11, TSHZ1, THY1, SEPT9, S100A4, CCDC92, TNS3, ENOX1, TGFB1I1, ZEB2, LMCD1, PDGFC, ECM1, TFPI, TBX3, DDR2, PTGER2, CDKN2C, VCAN, CD44, STARD13, SNED1, ZBTB38, SDC2, TPM1, COPZ2, STC1, CDH1, KRT5, KRT6B, CD24, EPCAM, ESRP1, CLDN3, CLDN4, ERBB3, SPOCK1, FERMT2, GLYR1, LTBP1, FADS2, KANK2, PTGFR, ARTN, SLC7A5, EPB41L4B, KRT15, CDH3, TPD52L1, ANXA3, PI3, NDRG1, DSG3, S100A14, S100S7, KLK8, S100A8, PTX3, SPINT2, FXYD3, KRT16, RAB25, LGALS7, DSC3, COL5A2, ITGB4, TMEM40, SPRR1A, AP1M2, SERPINB13, TP73L; and
- (d) CYBRD1, ARTN, CITED2, SLC7A5, CHN1, CTGF, CSTA, MYL9, EPB41L4B, KRT15, LRIG1, CDH3, TPD52L1, ANXA3, OLFML3, CORO1A, PI3, NDRG1, DSG3, PMP22, S100A14, S100S7, KLK8, S100A8, PTX3, SPINT2, IER3, FXYD3, KRT16, TGFBR3, KRT5, RAB25, SDC2, LUM, LGALS7, KRT6B, CD24, COL1A2, GNG11, DSC3, COL5A2, RGS4, ITGB4, TMEM40, SPRR1A, AP1M2, SERPINB13, SNED1, LEPREL1, TRAM2, TP73L, and PAP2B; and
- (e) CYBRD1, ARTN, CITED2, SLC7A5, CHN1, CTGF, CSTA, MYL9, EPB41L4B, KRT15, LRIG1, CDH3, TPD52L1, ANXA3, OLFML3, CORO1A; and
- (f) PI3, NDRG1, DSG3, PMP22, S100A14, S100S7, KLK8, S100A8, PTX3, SPINT2, IER3, FXYD3, KRT16, TGFBR3; and
- (g) KRT5, RAB25, SDC2, LUM, LGALS7, KRT6B, CD24, COL1A2, GNG11, DSC3, COL5A2; and
- (h) RGS4, ITGB4, TMEM40, SPRR1A, AP1M2, AERPINB13, SNED1, LEPREL1, TRAM2, TP73L, and PAB2B; and
  - (i) CYBRD1, ARTN, KRT15, ITGB4, RGS4, SPRR1B, SLC7A5, ANXA3, IL18, AP1M2, CHN1, OLFML3, ZBED2, LEPREL1, ABLIM1, CDH3, DNAJB4, TRAM2, UCHL1, CTGF; and
  - (i) CYBRD1, ARTN, KRT15, ITGB4, RGS4, SPRR1B, SLC7A5, ANXA3, IL18, AP1M2; and
  - (ii) CYBRD1, ARTN, KRT15, ITGB4, RGS4; and
- (j) CYBRD1, RGS4, CDH3, CHN1, SLC7A5, PRRX1, CITED2, COL5A2, ARTN, ZBED2, SNED1, ITGB4, LUM, PVRL3, OLFML3, BIN1, CCND2, DAB2, ANXA3, IL18; and
  - (i) CYBRD1, RGS4, CDH3, CHN1, SLC7A5, PRRX1, CITED2, COL5A2, ARTN, ZBED2; and
  - (ii) CYBRD1, RGS4, CDH3, CHN1, SLC7A5; and
- (k) CYBRD1, ARTN, RGS4, CITED2, SLC7A5, ITGB4, CHN1, PI3, TMEM40, NDRG1, SPRR1A, PCDH9, CTGF, MAP1B, DSG3, AP1M2, FBLN5, SERPINB13, PMP22, CSTA; and
  - (i) CYBRD1, ARTN, RGS4, CITED2, SLC7A5, ITGB4, CHN1, PI3, TMEM40, NDRG1; and
  - (ii) CYBRD1, ARTN, RGS4, CITED2, SLC7A5; and

(ii) said second set of genes comprises or consists of:
AXL, NOTCH3, GPR30, PROCR, PAX2, PI3KA, CD44, CD133, ALDH1, BMI1, KRT19, NRP1, MSI1, MTOR, JAG1, p38, ESA1, ZEB1, TWIST1, FOXC2, CDH2, ZEB2, FOXC1, SNAI1, SNAI2, TWIST2, GSC, TCF3, CD44, VIM, FN1, MLPH1, ARHGEF11; and
(iii) said third set of genes comprises or consists of:
At least one gene from said first set of genes and at least one of:
AXL, NOTCH3, GPR30, PROCR, PAX2, PI3KA, CD44, CD133, ALDH1, BMI1, KRT19, NRP1, MSI1, MTOR, JAG1, p38, ESA1, ZEB1, TWIST1, FOXC2, CDH2, ZEB2, FOXC1, SNAI1, SNAI2, TWIST2, GSC, TCF3, CD44, VIM, FN1, MLPH1, ARHGEF11; and
(iv) said fourth set of genes comprises or consists of:
At least two genes from said first set of genes and at least one of: AXL, NOTCH3, GPR30, PROCR, PAX2, PI3KA, CD44, CD133, ALDH1, BMI1, KRT19, NRP1, MSI1, MTOR, JAG1, p38, ESA1, ZEB1, TWIST1, FOXC2, CDH2, ZEB2, FOXC1, SNAI1, SNAI2, TWIST2, GSC, TCF3, CD44, VIM, FN1, MLPH1, and ARHGEF11; and
(v) said fifth set of genes comprises or consists of:
beta catenin, FZD1, FZD2, FZD3, FZD4, FZD5, FZD6, FZD7, FZD8, FZD9, FZD10, LRP5, LRP6, WNT1, WNT2, WNT2B, WNT3, WNT3A, WNT4 WNT5A, WNT5B, WNT6, WNT7A, WNT7B, WNT8A, WNT8B, WNT9A, WNT9B, WNT10A, WNT10B, WNT11, WNT16, SRFP1, SRFP2, FRZB, SFRP4, SFRP5, DKK1, DKK2, DKK3, DKK4, WIF1, AKT2, PI3KA, Disheveled, Smoothened, Axin1, Axin2; and
(vi) said sixth set of genes comprises or consists of:
At least one gene from said first set of genes and at least one of:
beta catenin, FZD1, FZD2, FZD3, FZD4, FZD5, FZD6, FZD7, FZD8, FZD9, FZD10, LRP5, LRP6, WNT1, WNT2, WNT2B, WNT3, WNT3A, WNT4 WNT5A, WNT5B, WNT6, WNT7A, WNT7B, WNT8A, WNT8B, WNT9A, WNT9B, WNT10A, WNT10B, WNT11, WNT16, SRFP1, SRFP2, FRZB, SFRP4, SFRP5, DKK1, DKK2, DKK3, DKK4, WIF1, AKT2, PI3KA, Disheveled, Smoothened, Axin1, Axin2; and
(vii) said seventh set of genes comprises or consists of:
TGFbeta1, TGFbeta2, TGFbeta3, ALK1, ALK2, ALK3, ALK4, ALK5, ALK6, ALK7, TGFbetaR-II, BMPR-II, ACTR-IIA, ACTR-IIB, AMHR-II, SMAD2, SMAD3; and
(viii) said eighth set of genes comprises or consists of:
At least one gene from said first set of genes and at least one of:
TGFbeta1, TGFbeta2, TGFbeta3, ALK1, ALK2, ALK3, ALK4, ALK5, ALK6, ALK7, TGFbetaR-II, BMPR-II, ACTR-IIA, ACTR-IIB, AMHR-II, SMAD2, SMAD3; and
(ix) said ninth set of genes comprises or consists of:
S100A4, SDC2, CYP1B1, FERMT2, CYBRD1, SNAI2, CD44, BIN1, PMP22, CTGF, IGFBP3, ANXA6, MAP1B, TAGLN, ENPP2, FBLN5, PCOLCE, FN1, VIM, PPAP2B, SEM5A, VCAN, NID2, NID1, HAS2, PRRX1, TNFAIP6, CLDN4, FADS2, DDIT4, GNG11, PER1, ISCU, KPNA3, SLC44A1, FKBP5, NR2F1, MLPH, DOCK10, RGS4, IGFBP4; and
(x) said tenth set of genes comprises or consists of:
S100A4, SDC2, CYP1B1, FERMT2, CYBRD1, SNAI2, CD44, BIN1, ENPP2, FBLN5, PCOLCE, FN1, VIM, PPAP2B, SEM5A, VCAN, CLDN4, FADS2, DDIT4, GNG11, PER1, ISCU, KPNA3, SLC44A1, FKBP5, IGFBP4; and
(xi) said eleventh set of genes comprises or consists of:
SDC2, CYBRD1, SNAI2, PMP22, CTGF, IGFBP3, ANXA6, MAP1B, TAGLN, PPAP2B, HAS2, NRF2, MLPH, DOCK10, RGS4; and
(xii) said twelfth set of genes comprises or consists of:
FERMT2, BIN1, PMP22, CHN1, CTGF, IGFBP3, NID2, CLDN4, CYBRD1, PRR16, ANXA6, PCOLCE, PPAP2B, HAS2, CDH11, MLPH, DOCK10, RGS4, PVRL3, NID2, SLC44A1, FKBP5; and
(xiii) said thirteenth set of genes comprises or consists of:
FERMT2, BIN1, PMP22, CHN1, CTGF, IGFBP3, NID2, CLDN4; and
(xiv) said fourteenth set of genes comprises or consists of:
HAS2, BIN1, PCOLCE, FERMT2, CTGF, IGFBP3, NID2, SLC44A1, FKBP5, and MLPH.
In an embodiment, said kit or product features a second agent capable of interacting with a gene expression product from said first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or seventh and/or eighth and/or ninth and/or tenth and/or eleventh and/or twelfth and/or thirteenth and/or fourteenth set of genes. In an embodiment, said kit or product features a plurality of agents capable of interacting with a gene expression product from said first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or seventh and/or eighth and/or ninth and/or tenth and/or eleventh and/or twelfth and/or thirteenth and/or fourteenth set of genes. In an embodiment, said kit or product features a plurality of agents capable of interacting with a plurality of gene expression products from said first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or seventh and/or eighth and/or ninth and/or tenth and/or eleventh and/or twelfth and/or thirteenth and/or fourteenth set of genes. In an embodiment, said plurality of gene expression products is any subset of gene expression products s in gene sets one, two, three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, or fourteen. In an embodiment, said plurality of gene expression products s is any subset of gene expression products in gene set one. In an embodiment, said plurality of gene expression products s is any subset of gene expression products in gene set two. In an embodiment, said plurality of gene expression products s is any subset of gene expression products in gene set three. In an embodiment, said plurality of gene expression products s is any subset of gene expression products in gene set four. In an embodiment, said plurality of gene expression products s is any subset of gene expression products in gene set five. In an embodiment, said plurality of gene expression products s is any subset of gene expression products in gene set six. In an embodiment, said plurality of gene expression products s is any subset of gene expression products in gene set seven. In an embodiment, said plurality of gene expression products s is any subset of gene expression products in gene set eight. In an embodiment, said plurality of gene expression products s is any subset of gene expression products in gene set nine. In an embodiment, said plurality of gene expression products s is any subset of gene expression products in gene set ten. In an embodiment, said plurality of gene expression products s is any subset of gene expression products in gene set eleven. In an embodiment, said plurality of gene expression products s is any subset of gene expression products in gene set twelve. In an embodiment, said plurality of gene expression products s is any subset of gene expression products in gene set thirteen. In an embodiment, said plurality of gene expression products s is any subset of gene expression products in gene set fourteen.
In an embodiment, said agent is a plurality of antibodies. In an embodiment, said agent is a plurality of oligonucleotides. In an embodiment, said agent is a plurality of antibodies and oligonucleotides. In an embodiment, said gene expression product is a RNA product. In an embodiment, said gene expression product is a protein product.
In an embodiment, said kit or product features an agent capable of interacting with a gene expression product of a gene in Table 1. In an embodiment, said kit or product contains plurality of agents capable of interacting with a plurality of genes in Table 1. In an embodiment, said kit or product features an agent capable of interacting with a gene expression product of a gene not in said first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or seventh and/or eighth and/or ninth and/or tenth and/or eleventh and/or twelfth and/or thirteenth and/or fourteenth set of genes. In an embodiment, said kit or product features a plurality of agents capable of interacting with a gene expression product of a plurality of genes not in said first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or seventh and/or eighth and/or ninth and/or tenth and/or eleventh and/or twelfth and/or thirteenth and/or fourteenth set of genes.
In one aspect, kits or products described herein include kits or products comprising a first agent capable of interacting with a gene expression product of a plurality of genes from a first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or seventh and/or eighth and/or ninth and/or tenth and/or eleventh and/or twelfth and/or thirteenth and/or fourteenth set of genes, wherein:
(i) said first set of genes comprises or consists of:

- KLF13, ANTXR2, IFNAR1, LIX1L, CHST11, AKAP2, PALM2-AKAPT, LOC80298, C7 orf2, NUCKS, DKFZP566D1346, LOC388279, FLJ31795, 6orf107, FLJ12439, ENPP2, CITED2, ZEB1, NID2, SEMA5A, DAB2, KCNMA1, PTX3, PCDH9, CREB3L1, BGN, ROR1, DLC1, GNG11, CDH11, NR2F1, PRR16, MYL9, DOCK10, LRIG1, IER3, EML1, NEBL, RGL1, MLPH, DNAJB4, FBLN5, RGS4, HAS2, ITGBL1, IGFBP4, DPT, PCOLCE, GREM1, PPAP2B, CDH2, PMP22, LUM, CHN1, CYP1B1, MME, WNT5A, POSTN, MMP2, CTGF, FBLN1, IGFBP3, DCN, PRRX1, ANXA6, PVRL3, MAP1B, TNFAIP6, CYBRD1, FBN1, NID1, OLFML3, SNAI1, SNAI2, TAGLN, SYNC, TUBA1A, PPM1D, TWIST1, FN1, TGFBR3, SERPINF1, UGDH, SRGN, FAP, PTGER4, PRKCA, FSTL1, MMP1, NRP1, FILIP1L, SCCPDH, LTBP2, XYLT1, HS3ST2, SYT11, TSHZ1, THY1, Sep9, S100A4, TNS3, ENOX1, TGFB1I1, ZEB2, LMCD1, PDGFC, ECM1, TFPI, TBX3, DDR2, PTGER2, CDKN2C, VCAN, CD44, STARD13, SNED1, ZBTB38, SDC2, TPM1, COPZ2, STC1, CDH1, KRT5, KRT6B, CD24, EPCAM, ESRP1, CLDN3, CLDN4, ERBB3, SPOCK1, FERMT2, GLYR1, LTBP1, FADS2, KANK2, PTGFR, COL11A2, KLK3, EIF2C2, ZFP41, FAM49B, PSORS1C2, MRPL42, MRPL54, MRPL47, MRPS23, EIF3S9, ALG5, DNAJC19, TPRXL, NOTCH2, RBM15, ST3GAL3, NFYA, PCNX, FBXO21, WWOX, CAMK2B, PNPLA2, CLIC5, UGCGL1, FBXL18, ADRBK1, SLC38A2, IL8RA, TAS2R14, CD300LB, GIPC3, RAD51L1, EPPK1, COL1A1, MMP9, SERPINE1, SPARC, TGFB1, TGFB3, TGFBI, TGFBR1, TGIF, TGIF2, THBS1, ANXA5, ACTG1, ARF3, ATP1B3, BAT3, CALD1, CENTD2, CLIC1, CTBS, DPYSL3, DVL3, EXT1, FGFR1, FTL, GNB2L1, GPRC5A, H2AFZ, HIF1A, IL13RA1, KDELR2, LARP1, LPIN2, MARS, MMP10, MMP14, MT2A, MYO10, NUP62, PFKFB3, PLOD2, PSMB7, PSMD8, RIN2, RYBP, SDF4, SETD5, SPP1, STAU1, TUBB3, UBE2S, XPNPEP1, CDKN1A, CHRD, H19, ID3, ID4, IGFBP7, LRP1, MSX1, NOTCH3, PROCR, GBX2, KI67, CCNB1, BUB1, KNTC2, USP22, HCFC1, RNF2, ANK3, FGFR2, CES1, COL1A2, COL3A1, COL5A2, COL6A1, ANKRD25, C10ORF56, C5ORF13, KRT81, NPAC, PLEKHC1, SEPT9, SYNC1, MBP, ABLIM1, ALDH1A3, ALOX15B, ANXA3, AP1M2, ARTN, CA2, CA9, CDH3, CDS1, COL17A1, CORO1A, CTSL2, CXADR, CYP27B1, DSC2, DSG3, DST, EPB41L4B, FGFBP1, FGFR3, FST, GJB3, GRHL2, HBEGF, HOOK1, IL18, IL1B, IRF6, ITGB4, JAG2, KLK10, KLK5, KLK7, KLK8, KRT15, KRT16, KRT17, LEPREL1, MYO5C, NDRG1, NMU, PI3, RAB25, RLN2, RNF128, S100A14, S100A7, S100A8, SERPINB1, SERPINB2, SLC2A9, SLPI, SNX10, SPINT2, SPRR1A, SPRR1B, ST14, SYK, TMEM40, TRIM29, TSPAN1, UCHL1, VSNL1, ZBED2, AQP3, CBARA1, CLCA2, CNTNAP2, CSTA, DSC3, ESRP2, FXYD3, GJB5, GPR87, IL1A, LAMA3, LAMB3, MAP7, MAPK13, MPZL2, NRG1, SCNN1A, SERPINB13, SLC7A5, SPINT1, ST6GALNAC2, TGFA, TP63, TPD52L1, VGLL1, SLC6A8, ABCA12, ANXA8, ANXA8L1, ARHGAP25, ARHGAP8, BDKRB2, BIK, C10ORF10, C10ORF116, C1ORF116, C20ORF19, C6ORF105, CELSR2, CKMT1A, CKMT1B, CLDN1, CST6, CYP4F11, E2F5, ELMO3, EPHA1, EVA1, EXPH5, F11R, FAT2, FLJ12684, FLJ20366, FZD3, GLS2, GNAL, IFI30, IGFBP2, IL1RN, IL4R, IRX4, JUP, KCNK1, KIAA0040, KIAA0888, KIAA1815, KRT14, KRT18, LAD1, LGALS7, LOC553158, LOC653562, LOC728113, LOC729884, LRRC1, LSR, MST1R, MYO1D, NAIP, OCLN, NEFM, NUP62CL, PERP, PLS1, POLR3G, PRKCH, PRRG4, PRSS8, PTPN3, RBM35A, RBM35B, RHBDF2, RHOD, RPS6KA1, RTEL1, SAA1, SAA1, SAA2, SH2D3A, SLC6A10P, SMPDL3B, SNCA, STAC, STAP2, TACSTD1, THBD, TMEM30B, TMPRSS11E, TMPRSS4, TNFRSF6B, TP73L, WWC1, XDH, ZNF165, AXL, GPR30, PAX2, PI3KA, CD133, ALDH1, BMI1, KRT19, MSI1, JAG1, p38, ESA1, FOXC2, FOXC1, TWIST2, GSC, TCF3, MLPH1, ARHGEF11a, DPF2, CASP8, BCL2, SCGN, SWAP70, KIAA0276, C10orf9, C10orf7, ALKBH, TOB2, XPR1, CD59, LRP2, PLP2, MAPK14, CXCL2, MMP7, MGP, MLF1, FLNB, SCNM1, HSPC163, C5orf18, MGC4399, CDW92, TMC4, ZDHHC2, TICAM2, KDELR3, GNPDA1, THEM2, DBR1, FLJ90709, FLJ10774, C16orf33, GAPD, LDHA, MR-1, LARS, GTPBP1, PRSS16, WFDC2, AIM1, DHRS6, DHRS4, GC15429, MGC45840, ECHDC2, GOLGIN-67, AFURS1, KIAA0436, CYP4V2, JTV1, ICMT, DNMT3A, HNMT, METTL7A, METTL2, VIL2, TPD52, ARPC5, NOL8, NSF, RAD23B, SRP54, HSPA2, PBP, THAP2, CIRBP, SNRPN, KIAA0052, DUSP10, SSR1, ERBB4, EMP1, CHPT1, LRPAP1, FLJ11752, CSTF1, KLHL20, DNAJC13, APLP2, ARGBP2, DNAJB1, NEBL, SH3BGRL, NUDT5, GABARAPL1, MAPT, DCBLD1, STK39, PAK2, CSNK2A1, PILRB, ERN1, SGKL, WEE1, MAST4, C11orf17, NUP37, CLTC, COPB2, SLC25A25, ECOP, PDE8A, STAM, TUBB, SNX6, RAB23, PLAA, STC2, LTF, ISGF3G, ATXN3, GTF3C3, GSK3B, KLF10, ELL2, ZBTB20, IRX3, ETS1, SERTAD1, MGC4251, MAFF, SFPQ, CITED4, CEBPD, EIF4E2, HS2ST1, AGPS, PGK1, ATIC, ETNK1, LG2, NCE2, MARCH8, CNOT4, RNF8, PSMA5, DPF2, AMMECR1, KIAA1287, LOC144233, LOC286505, PNAS-4, FLJ20530, HUMPD3, GC45564, CAP350, ETAA16, HAN11, DNAPTP6, C7orf25, FLJ37953, FLJ10587, C7orf36, ELP4, NDEL1, NPD014, KFZP564D172, FAM53C, IER5, LOC255783, KIAA0146, KIAA0792, LOC439994, LOC283481, CG018, LOC130576, NGFRAP1L1, KIAA1217, 4orf7, C21 orf86, C9orf64, FLJ13456, FLJ12806, FLJ39370, GATS, CCDC92, FMNL2, ARID1B, ZFHX1B, SSBP2, ARID5B, LOC157381, KPNA3, ARHGAP24, CCND2, VIM, MYCBP2, GAS7, TRAM2, BASP1, FOXO1A, POLR2A, PER1, DDIT4, CD97, BIN1, SH2B3, DDB2, EMP3, NDST2, CHST2, NT5E, PDE4A, CPS1, PTGS1, GGCX, IRF5, ZBTB16, MAP4K4, CHST7, KLF12, NFRKB, PSD, FKSG49, NIFUN, FYN, ZMYM2, CACNA1G, SLC25A16, FLII, EIF1, SEPT6, NPAC, PHF15, NUP188, ABR, CNR1, LOC283824, FSTL4, DNM1, APOBEC3G, ATP2B1, SMPD1, SLC11A1, FXYD5, C14orf139, SH3BGRL3, LUZP1, ZNF335, SH3 KBP1, MST150, PRO1073, LOC388397, FKBP5, HIPK2, DTX1, ST3GAL2, ADAMTS7, TNRC6B, CYGB, SDHAL1, LOC572558, TRIO, FRAS1, KIAA1632, POLS, EBF, MAML2, PTPRA, PLEKHG2, DYM, SOX6, ARHGEF2, ZCCHC6, PPP3CA, FAM70B, TMED5, FLJ43663, HPS1, MEF2A, ST3GAL5, SMYD3, KLF7, LOC200230, RERE, QKI, BICD1, CTNNB1, POU2F2, EIF4ENIF1, BTG1, TCHP, FLJ90709, PCTK2, KIAA1600, B7-H4, PDE4DIP, KIAA0194, HOM-TES; and
- (a) BGN, BIN1, CDH11, CDH2, CHN1, COL1A2, COL3A1, COL5A2, CREB3L1, CTGF, CYBRD1, DCN, DLC1, DNAJB4, DPT, ENPP2, FBLN1, FBLN5, FBN1, GREM1, HAS2, IGFBP3, IGFBP4, MLPH, MME, MMP2, MYL9, NEBL, NID1, NR2F1, OLFML3, PCOLCE, PMP22, POSTN, PPAP2B, PRR16, PRRX1, PTX3, PVRL3, RGL1, RGS4, ROR1, SEMA5A, TNFAIP6, TRAM2, WNT5A, ZEB1, ANXA6, CITED2, COL1A1, COL6A1, CYP1B1, DAB2, DOCK10, EML1, GNG11, IER3, ITGBL1, KCNMA1, LRIG1, LUM, MAP1B, NID2, PCDH9, PRKCA, PTGER4, PTGFR, SDC2, SERPINF1, SNED1, SYT11, TFPI, TGFBR3, VCAN, ANKRD25, C10ORF56, C5ORF13, CCDC92, CDKN2C, COPZ2, DDR2, ECM1, ENOX1, FADS2, FAP, FILIP1L, FSTL1, HS3ST2, KRT81, LMCD1, LTBP1, LTBP2, MMP1, NPAC, NRP1, PDGFC, PLEKHC1, PPM1D, PTGER2, SCCPDH, SEPT9, SPOCK1, SRGN, STARD13, STC1, SYNC1, TAGLN, TBX3, TGFB1I1, THY1, TNS3, TPM1, TSHZ1, TUBA1A, UGDH, VIM, XYLT1, ZBTB38, MBP, ABLIM1, ALDH1A3, ALOX15B, ANXA3, AP1M2, ARTN, CA2, CA9, CAMK2B, CCND2, CDH1, CDH3, CDS1, COL17A1, CORO1A, CTSL2, CXADR, CYP27B1, DSC2, DSG3, DST, EPB41L4B, FGFBP1, FGFR3, FST, GJB3, GRHL2, HBEGF, HOOK1, IL18, IL1B, IRF6, ITGB4, JAG2, KLK10, KLK5, KLK7, KLK8, KRT15, KRT16, KRT17, KRT5, KRT6B, LEPREL1, MYO5C, NDRG1, NMU, PI3, RAB25, RLN2, RNF128, S100A14, S100A7, S100A8, SERPINB1, SERPINB2, SLC2A9, SLPI, SNX10, SPINT2, SPRR1A, SPRR1B, ST14, SYK, TMEM40, TRIM29, TSPAN1, UCHL1, VSNL1, ZBED2, AQP3, CBARA1, CD24, CLCA2, CNTNAP2, CSTA, DSC3, EPCAM, ESRP2, FXYD3, GJB5, GPR87, IL1A, LAMA3, LAMB3, MAP7, MAPK13, MMP9, MPZL2, NRG1, SCNN1A, SERPINB13, SLC7A5, SPINT1, ST6GALNAC2, TGFA, TP63, TPD52L1, VGLL1, SLC6A8, ABCA12, ANK3, ANXA8, ANXA8L1, ARHGAP25, ARHGAP8, BDKRB2, BIK, C10ORF10, C10ORF116, C1ORF116, C20ORF19, C6ORF105, CELSR2, CKMT1A, CKMT1B, CLDN1, CST6, CYP4F11, E2F5, ELMO3, EPHA1, EVA1, EXPH5, F11R, FAT2, FGFR2, FLJ12684, FLJ20366, FZD3, GLS2, GNAL, IFI30, IGFBP2, IL1RN, IL4R, IRX4, JUP, KCNK1, KIAA0040, KIAA0888, KIAA1815, KRT14, KRT18, LAD1, LGALS7S, LOC553158, LOC653562, LOC728113, LOC729884, LRRC1, LSR, MST1R, MYO1D, NAIP, OCLN, NEFM, NUP62CL, PERP, PLS1, POLR3G, PRKCH, PRRG4, PRSS8, PTPN3, RBM35A, RBM35B, RHBDF2, RHOD, RPS6KA1, RTEL1, SAA1, SAA1, SAA2, SH2D3A, SLC6A10P, SMPDL3B, SNCA, STAC, STAP2, TACSTD1, THBD, TMEM30B, TMPRSS11E, TMPRSS4, TNFRSF6B, TP73L, WWC1, XDH, ZNF165; and
- (b) BGN, BIN1, CDH11, CDH2, CHN1, COL1A2, COL3A1, COL5A2, CREB3L1, CTGF, CYBRD1, DCN, DLC1, DNAJB4, DPT, ENPP2, FBLN1, FBLN5, FBN1, GREM1, HAS2, IGFBP3, IGFBP4, MLPH, MME, MMP2, MYL9, NEBL, NID1, NR2F1, OLFML3, PCOLCE, PMP22, POSTN, PPAP2B, PRR16, PRRX1, PTX3, PVRL3, RGL1, RGS4, ROR1, SEMA5A, TNFAIP6, TRAM2, WNT5A, ZEB1, ANXA6, CITED2, COL1A1, COL6A1, CYP1B1, DAB2, DOCK10, EML1, GNG11, IER3, ITGBL1, KCNMA1, LRIG1, LUM, MAP1B, NID2, PCDH9, PRKCA, PTGER4, PTGFR, SDC2, SERPINF1, SNED1, SYT11, TFPI, TGFBR3, VCAN, ANKRD25, C10ORF56, C5ORF13, CCDC92, CDKN2C, COPZ2, DDR2, ECM1, ENOX1, FADS2, FAP, FILIP1L, FSTL1, HS3ST2, KRT81, LMCD1, LTBP1, LTBP2, MMP1, NPAC, NRP1, PDGFC, PLEKHC1, PPM1D, PTGER2, SCCPDH, SEPT9, SPOCK1, SRGN, STARD13, STC1, SYNC1, TAGLN, TBX3, TGFB1I1, THY1, TNS3, TPM1, TSHZ1, TUBA1A, UGDH, VIM, XYLT1, ZBTB38; and
- (c) ENPP2, CITED2, ZEB1, NID2, SEMA5A, DAB2, KCNMA1, PTX3, PCDH9, CREB3L1, BGN, ROR1, DLC1, GNG11, CDH11, NR2F1, NR2F2, PRR16, MYL9, DOCK10, LRIG1, IER3, EML1, NEBL, RGL1, MLPH, TRAM2, DNAJB4, FBLN5, RGS4, HAS2, ITGBL1, IGFBP4, BIN1, DPT, PCOLCE, GREM1, PPAP2B, CDH2, PMP22, LUM, CHN1, CYP1B1, MME, WNT5A, POSTN, MMP2, CTGF, FBLN1, IGFBP3, DCN, PRRX1, ANXA6, PVRL3, MAP1B, TNFAIP6, CYBRD1, FBN1, NID1, OLFML3, SNAI1, SNAI2, TAGLN, SYNC, TUBA1A, PPM1D, TWIST1, FN1, TGFBR3, SERPINF1, UGDH, SRGN, FAP, PTGER4, VIM, PRKCA, FSTL1, MMP1, NRP1, FILIP1L, SCCPDH, LTBP2, XYLT1, HS3ST2, SYT11, TSHZ1, THY1, SEPT9, S100A4, CCDC92, TNS3, ENOX1, TGFB1I1, ZEB2, LMCD1, PDGFC, ECM1, TFPI, TBX3, DDR2, PTGER2, CDKN2C, VCAN, CD44, STARD13, SNED1, ZBTB38, SDC2, TPM1, COPZ2, STC1, CDH1, KRT5, KRT6B, CD24, EPCAM, ESRP1, CLDN3, CLDN4, ERBB3, SPOCK1, FERMT2, GLYR1, LTBP1, FADS2, KANK2, PTGFR, ARTN, SLC7A5, EPB41L4B, KRT15, CDH3, TPD52L1, ANXA3, PI3, NDRG1, DSG3, S100A14, S100S7, KLK8, S100A8, PTX3, SPINT2, FXYD3, KRT16, RAB25, LGALS7, DSG3, COL5A2, ITGB4, TMEM40, SPRR1A, AP1M2, SERPINB13, TP73L; and
- (d) CYBRD1, ARTN, CITED2, SLC7A5, CHN1, CTGF, CSTA, MYL9, EPB41L4B, KRT15, LRIG1, CDH3, TPD52L1, ANXA3, OLFML3, CORO1A, PI3, NDRG1, DSG3, PMP22, S100A14, S100S7, KLK8, S100A8, PTX3, SPINT2, IER3, FXYD3, KRT16, TGFBR3, KRT5, RAB25, SDC2, LUM, LGALS7, KRT6B, CD24, COL1A2, GNG11, DSC3, COL5A2, RGS4, ITGB4, TMEM40, SPRR1A, AP1M2, SERPINB13, SNED1, LEPREL1, TRAM2, TP73L, and PAP2B; and
- (e) CYBRD1, ARTN, CITED2, SLC7A5, CHN1, CTGF, CSTA, MYL9, EPB41L4B, KRT15, LRIG1, CDH3, TPD52L1, ANXA3, OLFML3, CORO1A; and
- (f) PI3, NDRG1, DSG3, PMP22, S100A14, S100S7, KLK8, S100A8, PTX3, SPINT2, IER3, FXYD3, KRT16, TGFBR3; and
- (g) KRT5, RAB25, SDC2, LUM, LGALS7, KRT6B, CD24, COL1A2, GNG11, DSC3, COL5A2; and
- (h) RGS4, ITGB4, TMEM40, SPRR1A, AP1M2, AERPINB13, SNED1, LEPREL1, TRAM2, TP73L, and PAB2B; and
- (i) CYBRD1, ARTN, KRT15, ITGB4, RGS4, SPRR1B, SLC7A5, ANXA3, IL18, AP1M2, CHN1, OLFML3, ZBED2, LEPREL1, ABLIM1, CDH3, DNAJB4, TRAM2, UCHL1, CTGF; and
  - (i) CYBRD1, ARTN, KRT15, ITGB4, RGS4, SPRR1B, SLC7A5, ANXA3, IL18, AP1M2; and
  - (ii) CYBRD1, ARTN, KRT15, ITGB4, RGS4; and
- (j) CYBRD1, RGS4, CDH3, CHN1, SLC7A5, PRRX1, CITED2, COL5A2, ARTN, ZBED2, SNED1, ITGB4, LUM, PVRL3, OLFML3, BIN1, CCND2, DAB2, ANXA3, IL18; and
  - (i) CYBRD1, RGS4, CDH3, CHN1, SLC7A5, PRRX1, CITED2, COL5A2, ARTN, ZBED2; and
  - (ii) CYBRD1, RGS4, CDH3, CHN1, SLC7A5; and
- (k) CYBRD1, ARTN, RGS4, CITED2, SLC7A5, ITGB4, CHN1, PI3, TMEM40, NDRG1, SPRR1A, PCDH9, CTGF, MAP1B, DSG3, AP1M2, FBLN5, SERPINB13, PMP22, CSTA; and
  - (i) CYBRD1, ARTN, RGS4, CITED2, SLC7A5, ITGB4, CHN1, PI3, TMEM40, NDRG1; and
  - (ii) CYBRD1, ARTN, RGS4, CITED2, SLC7A5; and

(ii) said second set of genes comprises or consists of:
AXL, NOTCH3, GPR30, PROCR, PAX2, PI3KA, CD44, CD133, ALDH1, BMI1, KRT19, NRP1, MSI1, MTOR, JAG1, p38, ESA1, ZEB1, TWIST1, FOXC2, CDH2, ZEB2, FOXC1, SNAI1, SNAI2, TWIST2, GSC, TCF3, CD44, VIM, FN1, MLPH1, ARHGEF11; and
(iii) said third set of genes comprises or consists of:
At least one gene from said first set of genes and at least one of:
AXL, NOTCH3, GPR30, PROCR, PAX2, PI3KA, CD44, CD133, ALDH1, BMI1, KRT19, NRP1, MSI1, MTOR, JAG1, p38, ESA1, ZEB1, TWIST1, FOXC2, CDH2, ZEB2, FOXC1, SNAI1, SNAI2, TWIST2, GSC, TCF3, CD44, VIM, FN1, MLPH1, ARHGEF11; and
(iv) said fourth set of genes comprises or consists of:
At least two genes from said first set of genes and at least one of: AXL, NOTCH3, GPR30, PROCR, PAX2, PI3KA, CD44, CD133, ALDH1, BMI1, KRT19, NRP1, MSI1, MTOR, JAG1, p38, ESA1, ZEB1, TWIST1, FOXC2, CDH2, ZEB2, FOXC1, SNAI1, SNAI2, TWIST2, GSC, TCF3, CD44, VIM, FN1, MLPH1, and ARHGEF11; and
(v) said fifth set of genes comprises or consists of:
beta catenin, FZD1, FZD2, FZD3, FZD4, FZD5, FZD6, FZD7, FZD8, FZD9, FZD10, LRP5, LRP6, WNT1, WNT2, WNT2B, WNT3, WNT3A, WNT4 WNT5A, WNT5B, WNT6, WNT7A, WNT7B, WNT8A, WNT8B, WNT9A, WNT9B, WNT10A, WNT10B, WNT11, WNT16, SRFP1, SRFP2, FRZB, SFRP4, SFRP5, DKK1, DKK2, DKK3, DKK4, WIF1, AKT2, PI3KA, Disheveled, Smoothened, Axin1, Axin2; and
(vi) said sixth set of genes comprises or consists of:
At least one gene from said first set of genes and at least one of:
beta catenin, FZD1, FZD2, FZD3, FZD4, FZD5, FZD6, FZD7, FZD8, FZD9, FZD10, LRP5, LRP6, WNT1, WNT2, WNT2B, WNT3, WNT3A, WNT4 WNT5A, WNT5B, WNT6, WNT7A, WNT7B, WNT8A, WNT8B, WNT9A, WNT9B, WNT10A, WNT10B, WNT11, WNT16, SRFP1, SRFP2, FRZB, SFRP4, SFRP5, DKK1, DKK2, DKK3, DKK4, WIF1, AKT2, PI3KA, Disheveled, Smoothened, Axin1, Axin2; and
(vii) said seventh set of genes comprises or consists of:
TGFbeta1, TGFbeta2, TGFbeta3, ALK1, ALK2, ALK3, ALK4, ALK5, ALK6, ALK7, TGFbetaR-II, BMPR-II, ACTR-IIA, ACTR-IIB, AMHR-II, SMAD2, SMAD3; and
(viii) said eighth set of genes comprises or consists of:
At least one gene from said first set of genes and at least one of:
TGFbeta1, TGFbeta2, TGFbeta3, ALK1, ALK2, ALK3, ALK4, ALK5, ALK6, ALK7, TGFbetaR-II, BMPR-II, ACTR-IIA, ACTR-IIB, AMHR-II, SMAD2, SMAD3; and
(ix) said ninth set of genes comprises or consists of:
S100A4, SDC2, CYP1B1, FERMT2, CYBRD1, SNAI2, CD44, BIN1, PMP22, CTGF, IGFBP3, ANXA6, MAP1B, TAGLN, ENPP2, FBLN5, PCOLCE, FN1, VIM, PPAP2B, SEM5A, VCAN, NID2, NID1, HAS2, PRRX1, TNFAIP6, CLDN4, FADS2, DDIT4, GNG11, PER1, ISCU, KPNA3, SLC44A1, FKBP5, NR2F1, MLPH, DOCK10, RGS4, IGFBP4; and
(x) said tenth set of genes comprises or consists of:
S100A4, SDC2, CYP1B1, FERMT2, CYBRD1, SNAI2, CD44, BIN1, ENPP2, FBLN5, PCOLCE, FN1, VIM, PPAP2B, SEM5A, VCAN, CLDN4, FADS2, DDIT4, GNG11, PER1, ISCU, KPNA3, SLC44A1, FKBP5, IGFBP4; and
(xi) said eleventh set of genes comprises or consists of:
SDC2, CYBRD1, SNAI2, PMP22, CTGF, IGFBP3, ANXA6, MAP1B, TAGLN, PPAP2B, HAS2, NRF2, MLPH, DOCK10, RGS4; and
(xii) said twelfth set of genes comprises or consists of:
FERMT2, BIN1, PMP22, CHN1, CTGF, IGFBP3, NID2, CLDN4, CYBRD1, PRR16, ANXA6, PCOLCE, PPAP2B, HAS2, CDH11, MLPH, DOCK10, RGS4, PVRL3, NID2, SLC44A1, FKBP5; and
(xiii) said thirteenth set of genes comprises or consists of:
FERMT2, BIN1, PMP22, CHN1, CTGF, IGFBP3, NID2, CLDN4; and
(xiv) said fourteenth set of genes comprises or consists of:
HAS2, BIN1, PCOLCE, FERMT2, CTGF, IGFBP3, NID2, SLC44A1, FKBP5, and MLPH.
In one embodiment, the kit or product comprises a second agent capable of interacting with a gene expression product from said first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or seventh and/or eighth and/or ninth and/or tenth and/or eleventh and/or twelfth and/or thirteenth and/or fourteenth set of genes. In one embodiment, the kit or product comprises a plurality of agents capable of interacting with a gene expression product from said first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or seventh and/or eighth and/or ninth and/or tenth and/or eleventh and/or twelfth and/or thirteenth and/or fourteenth set of genes. In one embodiment, the kit or product comprises a plurality of agents capable of interacting with a plurality of gene expression products from said first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or seventh and/or eighth and/or ninth and/or tenth and/or eleventh and/or twelfth and/or thirteenth and/or fourteenth set of genes.
In one embodiment, said agent is a plurality of antibodies. In one embodiment, said agent is a plurality of oligonucleotides. In one embodiment, said gene expression product is a RNA product. In one embodiment, said gene expression product is a protein product.
In one embodiment, the gene expression products are derived from a tumor sample, e.g., a preparation of a primary tumor, metastatic tumor, lymph node, circulating tumor cells, ascites, or pleural effusion, plasma, serum, circulating, and interstitial fluid levels.
In one embodiment, a value for the level of gene expression product for each gene is determined. In one embodiment, a value that is a function of the level of gene expression for each gene is determined. In one embodiment, the value is compared to a reference standard, e.g., the level of expression of a control gene in the tumor sample.
In one embodiment, the kit or product further comprises the performance of an algorithm on a computer system to determine a value or values that is a function of a location of a gene expression product in the subject sample and/or a function of a level of a gene expression product of a gene in the subject sample. In one embodiment, the algorithm compares a ratio of the level of gene expression product of at least one of the genes selected from the group: HAS2, BIN1, PCOLCE, FERMT2, CTGF, IGFBP3, NID2, SLC44A1, FKBP5, and MLPH; to the level of gene expression product of at least one of the genes selected from the group: CDH1, and Cytokeratin.
In one embodiment, the kit or product further comprises a plurality of agents capable of interacting with at least one gene expression product selected from the group: CTGF, IGFBP3, TNFAIP6, NID2, HAS2, CCL2, MLPH, NID1, IGFBP4, FBLN5, and PCOLCE. In one embodiment, the kit or product further comprises a plurality of agents capable of interacting with a gene expression product of each gene from the set of genes consisting of: CTGF, IGFBP3, TNFAIP6, NID2, HAS2, CCL2, MLPH, NID1, IGFBP4, FBLN5, and PCOLCE.
In one aspect, kits or products described herein include kits or products comprising a first agent capable of interacting with a gene expression product of a gene from a first and/or second and/or third and/or fourth and/or fifth and/or sixth set of genes, wherein:
(i) said first set of genes comprises or consists of:

- S100A4, SDC2, CYP1B1, FERMT2, CYBRD1, SNAI2, CD44, BIN1, PMP22, CTGF, IGFBP3, ANXA6, MAP1B, TAGLN, ENPP2, FBLN5, PCOLCE, FN1, VIM, PPAP2B, SEM5A, VCAN, NID2, NID1, HAS2, PRRX1, TNFAIP6, CLDN4, FADS2, DDIT4, GNG11, PER1, ISCU, KPNA3, SLC44A1, FKBP5, NR2F1, MLPH, DOCK10, RGS4, IGFBP4; and

(ii) said second set of genes comprises or consists of:

- S100A4, SDC2, CYP1B1, FERMT2, CYBRD1, SNAI2, CD44, BIN1, ENPP2, FBLN5, PCOLCE, FN1, VIM, PPAP2B, SEM5A, VCAN, CLDN4, FADS2, DDIT4, GNG11, PER1, ISCU, KPNA3, SLC44A1, FKBP5, IGFBP4; and

(iii) said third set of genes comprises or consists of:

- SDC2, CYBRD1, SNAI2, PMP22, CTGF, IGFBP3, ANXA6, MAP1B, TAGLN, PPAP2B, HAS2, NRF2, MLPH, DOCK10, RGS4; and

(iv) said fourth set of genes comprises or consists of:

- FERMT2, BIN1, PMP22, CHN1, CTGF, IGFBP3, NID2, CLDN4, CYBRD1, PRR16, ANXA6, PCOLCE, PPAP2B, HAS2, CDH11, MLPH, DOCK10, RGS4, PVRL3, NID2, SLC44A1, FKBP5; and

(v) said fifth set of genes comprises or consists of:

- FERMT2, BIN1, PMP22, CHN1, CTGF, IGFBP3, NID2, CLDN4; and

(vi) said sixth set of genes comprises or consists of:

- HAS2, BIN1, PCOLCE, FERMT2, CTGF, IGFBP3, NID2, SLC44A1, FKBP5, and MLPH.

In one embodiment, the kit or product comprises a second agent capable of interacting with a gene expression product of a plurality of genes from said first and/or second and/or third and/or fourth and/or fifth and/or sixth set of genes. In one embodiment, the kit or product comprises a plurality of agents capable of interacting with a gene expression product of a plurality of genes from said first and/or second and/or third and/or fourth and/or fifth and/or sixth set of genes. In one embodiment, the kit or product comprises a plurality of agents capable of interacting with a plurality of gene expression products of a plurality of genes from said first and/or second and/or third and/or fourth and/or fifth and/or sixth set of genes.
In one embodiment, said agent is a plurality of antibodies. In one embodiment, said agent is a plurality of oligonucleotides. In one embodiment, said gene expression product is a RNA product. In one embodiment, said gene expression product is a protein product. In one embodiment, a value for the level of gene expression product for each gene isoform is assayed. In one embodiment, a value for the level of gene expression product for each gene isoform is assayed by detecting a protein product. In one embodiment, the protein product is detected by an immunoassay, e.g., immunohistochemistry. In one embodiment, a value for the level of gene expression product for each gene isoform is assayed by detecting a RNA product. In one embodiment, the RNA product is detected by a hybridization based method. In one embodiment, the RNA product is detected by microarray. In one embodiment, said microarray is an exon microarray. In one embodiment, the RNA product is detected by a polymerase chain reaction based method. In one embodiment, the RNA product is detected by a sequencing based method. In one embodiment, the RNA product is detected by a quantitative RNA sequencing.
In one embodiment, the gene expression products are derived from a tumor sample, e.g., a preparation of a primary tumor, metastatic tumor, lymph node, circulating tumor cells, ascites, or pleural effusion, plasma, serum, circulating, and interstitial fluid levels.
In one embodiment, a value for the level of gene expression product for each gene is determined. In one embodiment, a value that is a function of the level of gene expression for each gene is determined. In one embodiment, the value is compared to a reference standard, e.g., the level of expression of a control gene in the tumor sample.
In one embodiment, the kit or product further comprises the performance of an algorithm on a computer system to determine a value or values that is a function of a location of a gene expression product in the subject sample and/or a function of a level of a gene expression product of a gene in the subject sample. In one embodiment, the algorithm compares a ratio of the level of gene expression product of at least one of the genes selected from the group: HAS2, BIN1, PCOLCE, FERMT2, CTGF, IGFBP3, NID2, SLC44A1, FKBP5, and MLPH; to the level of gene expression product of at least one of the genes selected from the group: CDH1, and Cytokeratin.
In one embodiment, the kit or product further comprises a plurality of agents capable of interacting with at least one gene expression product selected from the group: CTGF, IGFBP3, TNFAIP6, NID2, HAS2, CCL2, MLPH, NID1, IGFBP4, FBLN5, and PCOLCE. In one embodiment, the kit or product further comprises a plurality of agents capable of interacting with a gene expression product of each gene from the set of genes consisting of: CTGF, IGFBP3, TNFAIP6, NID2, HAS2, CCL2, MLPH, NID1, IGFBP4, FBLN5, and PCOLCE.
Methods of Assaying
In one aspect, methods described herein include methods of assaying in a subject sample the level of gene expression product of a plurality of genes from a first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or seventh and/or eighth and/or ninth and/or tenth and/or eleventh and/or twelfth and/or thirteenth and/or fourteenth set of genes, wherein:
(i) said first set of genes comprises or consists of:

- KLF13, ANTXR2, IFNAR1, LIX1L, CHST11, AKAP2, PALM2-AKAPT, LOC80298, C7orf2, NUCKS, DKFZP566D1346, LOC388279, FLJ31795, 6orf107, FLJ12439, ENPP2, CITED2, ZEB1, NID2, SEMA5A, DAB2, KCNMA1, PTX3, PCDH9, CREB3L1, BGN, ROR1, DLC1, GNG11, CDH11, NR2F1, PRR16, MYL9, DOCK10, LRIG1, IER3, EML1, NEBL, RGL1, MLPH, DNAJB4, FBLN5, RGS4, HAS2, ITGBL1, IGFBP4, DPT, PCOLCE, GREM1, PPAP2B, CDH2, PMP22, LUM, CHN1, CYP1B1, MME, WNT5A, POSTN, MMP2, CTGF, FBLN1, IGFBP3, DCN, PRRX1, ANXA6, PVRL3, MAP1B, TNFAIP6, CYBRD1, FBN1, NID1, OLFML3, SNAI1, SNAI2, TAGLN, SYNC, TUBA1A, PPM1D, TWIST1, FN1, TGFBR3, SERPINF1, UGDH, SRGN, FAP, PTGER4, PRKCA, FSTL1, MMP1, NRP1, FILIP1L, SCCPDH, LTBP2, XYLT1, HS3ST2, SYT11, TSHZ1, THY1, Sep9, S100A4, TNS3, ENOX1, TGFB1I1, ZEB2, LMCD1, PDGFC, ECM1, TFPI, TBX3, DDR2, PTGER2, CDKN2C, VCAN, CD44, STARD13, SNED1, ZBTB38, SDC2, TPM1, COPZ2, STC1, CDH1, KRT5, KRT6B, CD24, EPCAM, ESRP1, CLDN3, CLDN4, ERBB3, SPOCK1, FERMT2, GLYR1, LTBP1, FADS2, KANK2, PTGFR, COL11A2, KLK3, EIF2C2, ZFP41, FAM49B, PSORS1C2, MRPL42, MRPL54, MRPL47, MRPS23, EIF3S9, ALG5, DNAJC19, TPRXL, NOTCH2, RBM15, ST3GAL3, NFYA, PCNX, FBXO21, WWOX, CAMK2B, PNPLA2, CLIC5, UGCGL1, FBXL18, ADRBK1, SLC38A2, IL8RA, TAS2R14, CD300LB, GIPC3, RAD51L1, EPPK1, COL1A1, MMP9, SERPINE1, SPARC, TGFB1, TGFB3, TGFBI, TGFBR1, TGIF, TGIF2, THBS1, ANXA5, ACTG1, ARF3, ATP1B3, BAT3, CALD1, CENTD2, CLIC1, CTBS, DPYSL3, DVL3, EXT1, FGFR1, FTL, GNB2L1, GPRC5A, H2AFZ, HIF1A, IL13RA1, KDELR2, LARP1, LPIN2, MARS, MMP10, MMP14, MT2A, MYO10, NUP62, PFKFB3, PLOD2, PSMB7, PSMD8, RIN2, RYBP, SDF4, SETD5, SPP1, STAU1, TUBB3, UBE2S, XPNPEP1, CDKN1A, CHRD, H19, ID3, ID4, IGFBP7, LRP1, MSX1, NOTCH3, PROCR, GBX2, KI67, CCNB1, BUB1, KNTC2, USP22, HCFC1, RNF2, ANK3, FGFR2, CES1, COL1A2, COL3A1, COL5A2, COL6A1, ANKRD25, C10ORF56, C5ORF13, KRT81, NPAC, PLEKHC1, SEPT9, SYNC1, MBP, ABLIM1, ALDH1A3, ALOX15B, ANXA3, AP1M2, ARTN, CA2, CA9, CDH3, CDS1, COL17A1, CORO1A, CTSL2, CXADR, CYP27B1, DSC2, DSG3, DST, EPB41L4B, FGFBP1, FGFR3, FST, GJB3, GRHL2, HBEGF, HOOK1, IL18, IL1B, IRF6, ITGB4, JAG2, KLK10, KLK5, KLK7, KLK8, KRT15, KRT16, KRT17, LEPREL1, MYO5C, NDRG1, NMU, PI3, RAB25, RLN2, RNF128, S100A14, S100A7, S100A8, SERPINB1, SERPINB2, SLC2A9, SLPI, SNX10, SPINT2, SPRR1A, SPRR1B, ST14, SYK, TMEM40, TRIM29, TSPAN1, UCHL1, VSNL1, ZBED2, AQP3, CBARA1, CLCA2, CNTNAP2, CSTA, DSC3, ESRP2, FXYD3, GJB5, GPR87, IL1A, LAMA3, LAMB3, MAP7, MAPK13, MPZL2, NRG1, SCNN1A, SERPINB13, SLC7A5, SPINT1, ST6GALNAC2, TGFA, TP63, TPD52L1, VGLL1, SLC6A8, ABCA12, ANXA8, ANXA8L1, ARHGAP25, ARHGAP8, BDKRB2, BIK, C10ORF10, C10ORF116, C1ORF116, C20ORF19, C6ORF105, CELSR2, CKMT1A, CKMT1B, CLDN1, CST6, CYP4F11, E2F5, ELMO3, EPHA1, EVA1, EXPH5, F11R, FAT2, FLJ12684, FLJ20366, FZD3, GLS2, GNAL, IFI30, IGFBP2, IL1RN, IL4R, IRX4, JUP, KCNK1, KIAA0040, KIAA0888, KIAA1815, KRT14, KRT18, LAD1, LGALS7, LOC553158, LOC653562, LOC728113, LOC729884, LRRC1, LSR, MST1R, MYO1D, NAIP, OCLN, NEFM, NUP62CL, PERP, PLS1, POLR3G, PRKCH, PRRG4, PRSS8, PTPN3, RBM35A, RBM35B, RHBDF2, RHOD, RPS6KA1, RTEL1, SAA1, SAA1, SAA2, SH2D3A, SLC6A10P, SMPDL3B, SNCA, STAC, STAP2, TACSTD1, THBD, TMEM30B, TMPRSS11E, TMPRSS4, TNFRSF6B, TP73L, WWC1, XDH, ZNF165, AXL, GPR30, PAX2, PI3KA, CD133, ALDH1, BMI1, KRT19, MSI1, JAG1, p38, ESA1, FOXC2, FOXC1, TWIST2, GSC, TCF3, MLPH1, ARHGEF11a, DPF2, CASP8, BCL2, SCGN, SWAP70, KIAA0276, C10orf9, C10orf7, ALKBH, TOB2, XPR1, CD59, LRP2, PLP2, MAPK14, CXCL2, MMP7, MGP, MLF1, FLNB, SCNM1, HSPC163, C5orf18, MGC4399, CDW92, TMC4, ZDHHC2, TICAM2, KDELR3, GNPDA1, THEM2, DBR1, FLJ90709, FLJ10774, C16orf33, GAPD, LDHA, MR-1, LARS, GTPBP1, PRSS16, WFDC2, AIM1, DHRS6, DHRS4, GC15429, MGC45840, ECHDC2, GOLGIN-67, AFURS1, KIAA0436, CYP4V2, JTV1, ICMT, DNMT3A, HNMT, METTL7A, METTL2, VIL2, TPD52, ARPC5, NOL8, NSF, RAD23B, SRP54, HSPA2, PBP, THAP2, CIRBP, SNRPN, KIAA0052, DUSP10, SSR1, ERBB4, EMP1, CHPT1, LRPAP1, FLJ11752, CSTF1, KLHL20, DNAJC13, APLP2, ARGBP2, DNAJB1, NEBL, SH3BGRL, NUDT5, GABARAPL1, MAPT, DCBLD1, STK39, PAK2, CSNK2A1, PILRB, ERN1, SGKL, WEE1, MAST4, C11orf17, NUP37, CLTC, COPB2, SLC25A25, ECOP, PDE8A, STAM, TUBB, SNX6, RAB23, PLAA, STC2, LTF, ISGF3G, ATXN3, GTF3C3, GSK3B, KLF10, ELL2, ZBTB20, IRX3, ETS1, SERTAD1, MGC4251, MAFF, SFPQ, CITED4, CEBPD, EIF4E2, HS2ST1, AGPS, PGK1, ATIC, ETNK1, LG2, NCE2, MARCH8, CNOT4, RNF8, PSMA5, DPF2, AMMECR1, KIAA1287, LOC144233, LOC286505, PNAS-4, FLJ20530, HUMPD3, GC45564, CAP350, ETAA16, HAN11, DNAPTP6, C7orf25, FLJ37953, FLJ10587, C7orf36, ELP4, NDEL1, NPD014, KFZP564D172, FAM53C, IER5, LOC255783, KIAA0146, KIAA0792, LOC439994, LOC283481, CG018, LOC130576, NGFRAP1L1, KIAA1217, 4orf7, C21 orf86, C9orf64, FLJ13456, FLJ12806, FLJ39370, GATS, CCDC92, FMNL2, ARID1B, ZFHX1B, SSBP2, ARID5B, LOC157381, KPNA3, ARHGAP24, CCND2, VIM, MYCBP2, GAS7, TRAM2, BASP1, FOXO1A, POLR2A, PER1, DDIT4, CD97, BIN1, SH2B3, DDB2, EMP3, NDST2, CHST2, NT5E, PDE4A, CPS1, PTGS1, GGCX, IRF5, ZBTB16, MAP4K4, CHST7, KLF12, NFRKB, PSD, FKSG49, NIFUN, FYN, ZMYM2, CACNA1G, SLC25A16, FLII, EIF1, SEPT6, NPAC, PHF15, NUP188, ABR, CNR1, LOC283824, FSTL4, DNM1, APOBEC3G, ATP2B1, SMPD1, SLC11A1, FXYD5, C14orf139, SH3BGRL3, LUZP1, ZNF335, SH3 KBP1, MST150, PRO1073, LOC388397, FKBP5, HIPK2, DTX1, ST3GAL2, ADAMTS7, TNRC6B, CYGB, SDHAL1, LOC572558, TRIO, FRAS1, KIAA1632, POLS, EBF, MAML2, PTPRA, PLEKHG2, DYM, SOX6, ARHGEF2, ZCCHC6, PPP3CA, FAM70B, TMED5, FLJ43663, HPS1, MEF2A, ST3GAL5, SMYD3, KLF7, LOC200230, RERE, QKI, BICD1, CTNNB1, POU2F2, EIF4ENIF1, BTG1, TCHP, FLJ90709, PCTK2, KIAA1600, B7-H4, PDE4DIP, KIAA0194, HOM-TES; and
- (a) BGN, BIN1, CDH11, CDH2, CHN1, COL1A2, COL3A1, COL5A2, CREB3L1, CTGF, CYBRD1, DCN, DLC1, DNAJB4, DPT, ENPP2, FBLN1, FBLN5, FBN1, GREM1, HAS2, IGFBP3, IGFBP4, MLPH, MME, MMP2, MYL9, NEBL, NID1, NR2F1, OLFML3, PCOLCE, PMP22, POSTN, PPAP2B, PRR16, PRRX1, PTX3, PVRL3, RGL1, RGS4, ROR1, SEMA5A, TNFAIP6, TRAM2, WNT5A, ZEB1, ANXA6, CITED2, COL1A1, COL6A1, CYP1B1, DAB2, DOCK10, EML1, GNG11, IER3, ITGBL1, KCNMA1, LRIG1, LUM, MAP1B, NID2, PCDH9, PRKCA, PTGER4, PTGFR, SDC2, SERPINF1, SNED1, SYT11, TFPI, TGFBR3, VCAN, ANKRD25, C10ORF56, C5ORF13, CCDC92, CDKN2C, COPZ2, DDR2, ECM1, ENOX1, FADS2, FAP, FILIP1L, FSTL1, HS3ST2, KRT81, LMCD1, LTBP1, LTBP2, MMP1, NPAC, NRP1, PDGFC, PLEKHC1, PPM1D, PTGER2, SCCPDH, SEPT9, SPOCK1, SRGN, STARD13, STC1, SYNC1, TAGLN, TBX3, TGFB1I1, THY1, TNS3, TPM1, TSHZ1, TUBA1A, UGDH, VIM, XYLT1, ZBTB38, MBP, ABLIM1, ALDH1A3, ALOX15B, ANXA3, AP1M2, ARTN, CA2, CA9, CAMK2B, CCND2, CDH1, CDH3, CDS1, COL17A1, CORO1A, CTSL2, CXADR, CYP27B1, DSC2, DSG3, DST, EPB41L4B, FGFBP1, FGFR3, FST, GJB3, GRHL2, HBEGF, HOOK1, IL18, IL1B, IRF6, ITGB4, JAG2, KLK10, KLK5, KLK7, KLK8, KRT15, KRT16, KRT17, KRT5, KRT6B, LEPREL1, MYO5C, NDRG1, NMU, PI3, RAB25, RLN2, RNF128, S100A14, S100A7, S100A8, SERPINB1, SERPINB2, SLC2A9, SLPI, SNX10, SPINT2, SPRR1A, SPRR1B, ST14, SYK, TMEM40, TRIM29, TSPAN1, UCHL1, VSNL1, ZBED2, AQP3, CBARA1, CD24, CLCA2, CNTNAP2, CSTA, DSC3, EPCAM, ESRP2, FXYD3, GJB5, GPR87, IL1A, LAMA3, LAMB3, MAP7, MAPK13, MMP9, MPZL2, NRG1, SCNN1A, SERPINB13, SLC7A5, SPINT1, ST6GALNAC2, TGFA, TP63, TPD52L1, VGLL1, SLC6A8, ABCA12, ANK3, ANXA8, ANXA8L1, ARHGAP25, ARHGAP8, BDKRB2, BIK, C10ORF10, C10ORF116, C1ORF116, C20ORF19, C6ORF105, CELSR2, CKMT1A, CKMT1B, CLDN1, CST6, CYP4F11, E2F5, ELMO3, EPHA1, EVA1, EXPH5, F11R, FAT2, FGFR2, FLJ12684, FLJ20366, FZD3, GLS2, GNAL, IFI30, IGFBP2, IL1RN, IL4R, IRX4, JUP, KCNK1, KIAA0040, KIAA0888, KIAA1815, KRT14, KRT18, LAD1, LGALS7, LOC553158, LOC653562, LOC728113, LOC729884, LRRC1, LSR, MST1R, MYO1D, NAIP, OCLN, NEFM, NUP62CL, PERP, PLS1, POLR3G, PRKCH, PRRG4, PRSS8, PTPN3, RBM35A, RBM35B, RHBDF2, RHOD, RPS6KA1, RTEL1, SAA1, SAA1, SAA2, SH2D3A, SLC6A10P, SMPDL3B, SNCA, STAC, STAP2, TACSTD1, THBD, TMEM30B, TMPRSS11E, TMPRSS4, TNFRSF6B, TP73L, WWC1, XDH, ZNF165; and
- (b) BGN, BIN1, CDH11, CDH2, CHN1, COL1A2, COL3A1, COL5A2, CREB3L1, CTGF, CYBRD1, DCN, DLC1, DNAJB4, DPT, ENPP2, FBLN1, FBLN5, FBN1, GREM1, HAS2, IGFBP3, IGFBP4, MLPH, MME, MMP2, MYL9, NEBL, NID1, NR2F1, OLFML3, PCOLCE, PMP22, POSTN, PPAP2B, PRR16, PRRX1, PTX3, PVRL3, RGL1, RGS4, ROR1, SEMA5A, TNFAIP6, TRAM2, WNT5A, ZEB1, ANXA6, CITED2, COL1A1, COL6A1, CYP1B1, DAB2, DOCK10, EML1, GNG11, IER3, ITGBL1, KCNMA1, LRIG1, LUM, MAP1B, NID2, PCDH9, PRKCA, PTGER4, PTGFR, SDC2, SERPINF1, SNED1, SYT11, TFPI, TGFBR3, VCAN, ANKRD25, C10ORF56, C5ORF13, CCDC92, CDKN2C, COPZ2, DDR2, ECM1, ENOX1, FADS2, FAP, FILIP1L, FSTL1, HS3ST2, KRT81, LMCD1, LTBP1, LTBP2, MMP1, NPAC, NRP1, PDGFC, PLEKHC1, PPM1D, PTGER2, SCCPDH, SEPT9, SPOCK1, SRGN, STARD13, STC1, SYNC1, TAGLN, TBX3, TGFB1I1, THY1, TNS3, TPM1, TSHZ1, TUBA1A, UGDH, VIM, XYLT1, ZBTB38; and
- (c) ENPP2, CITED2, ZEB1, NID2, SEMA5A, DAB2, KCNMA1, PTX3, PCDH9, CREB3L1, BGN, ROR1, DLC1, GNG11, CDH11, NR2F1, NR2F2, PRR16, MYL9, DOCK10, LRIG1, IER3, EML1, NEBL, RGL1, MLPH, TRAM2, DNAJB4, FBLN5, RGS4, HAS2, ITGBL1, IGFBP4, BIN1, DPT, PCOLCE, GREM1, PPAP2B, CDH2, PMP22, LUM, CHN1, CYP1B1, MME, WNT5A, POSTN, MMP2, CTGF, FBLN1, IGFBP3, DCN, PRRX1, ANXA6, PVRL3, MAP1B, TNFAIP6, CYBRD1, FBN1, NID1, OLFML3, SNAI1, SNAI2, TAGLN, SYNC, TUBA1A, PPM1D, TWIST1, FN1, TGFBR3, SERPINF1, UGDH, SRGN, FAP, PTGER4, VIM, PRKCA, FSTL1, MMP1, NRP1, FILIP1L, SCCPDH, LTBP2, XYLT1, HS3ST2, SYT11, TSHZ1, THY1, SEPT9, S100A4, CCDC92, TNS3, ENOX1, TGFB1I1, ZEB2, LMCD1, PDGFC, ECM1, TFPI, TBX3, DDR2, PTGER2, CDKN2C, VCAN, CD44, STARD13, SNED1, ZBTB38, SDC2, TPM1, COPZ2, STC1, CDH1, KRT5, KRT6B, CD24, EPCAM, ESRP1, CLDN3, CLDN4, ERBB3, SPOCK1, FERMT2, GLYR1, LTBP1, FADS2, KANK2, PTGFR, ARTN, SLC7A5, EPB41L4B, KRT15, CDH3, TPD52L1, ANXA3, PI3, NDRG1, DSG3, S100A14, S100S7, KLK8, S100A8, PTX3, SPINT2, FXYD3, KRT16, RAB25, LGALS7, DSC3, COL5A2, ITGB4, TMEM40, SPRR1A, AP1M2, SERPINB13, TP73L; and
- (d) CYBRD1, ARTN, CITED2, SLC7A5, CHN1, CTGF, CSTA, MYL9, EPB41L4B, KRT15, LRIG1, CDH3, TPD52L1, ANXA3, OLFML3, CORO1A, PI3, NDRG1, DSG3, PMP22, S100A14, S100S7, KLK8, S100A8, PTX3, SPINT2, IER3, FXYD3, KRT16, TGFBR3, KRT5, RAB25, SDC2, LUM, LGALS7, KRT6B, CD24, COL1A2, GNG11, DSC3, COL5A2, RGS4, ITGB4, TMEM40, SPRR1A, AP1M2, SERPINB13, SNED1, LEPREL1, TRAM2, TP73L, and PAP2B; and
- (e) CYBRD1, ARTN, CITED2, SLC7A5, CHN1, CTGF, CSTA, MYL9, EPB41L4B, KRT15, LRIG1, CDH3, TPD52L1, ANXA3, OLFML3, CORO1A; and
- (f) PI3, NDRG1, DSG3, PMP22, S100A14, S100S7, KLK8, S100A8, PTX3, SPINT2, IER3, FXYD3, KRT16, TGFBR3; and
- (g) KRT5, RAB25, SDC2, LUM, LGALS7, KRT6B, CD24, COL1A2, GNG11, DSC3, COL5A2; and
- (h) RGS4, ITGB4, TMEM40, SPRR1A, AP1M2, AERPINB13, SNED1, LEPREL1, TRAM2, TP73L, and PAB2B; and
- (i) CYBRD1, ARTN, KRT15, ITGB4, RGS4, SPRR1B, SLC7A5, ANXA3, IL18, AP1M2, CHN1, OLFML3, ZBED2, LEPREL1, ABLIM1, CDH3, DNAJB4, TRAM2, UCHL1, CTGF; and
  - (i) CYBRD1, ARTN, KRT15, ITGB4, RGS4, SPRR1B, SLC7A5, ANXA3, IL18, AP1M2; and
  - (ii) CYBRD1, ARTN, KRT15, ITGB4, RGS4; and
- (j) CYBRD1, RGS4, CDH3, CHN1, SLC7A5, PRRX1, CITED2, COL5A2, ARTN, ZBED2, SNED1, ITGB4, LUM, PVRL3, OLFML3, BIN1, CCND2, DAB2, ANXA3, IL18; and
  - (i) CYBRD1, RGS4, CDH3, CHN1, SLC7A5, PRRX1, CITED2, COL5A2, ARTN, ZBED2; and
  - (ii) CYBRD1, RGS4, CDH3, CHN1, SLC7A5; and
- (k) CYBRD1, ARTN, RGS4, CITED2, SLC7A5, ITGB4, CHN1, PI3, TMEM40, NDRG1, SPRR1A, PCDH9, CTGF, MAP1B, DSG3, AP1M2, FBLN5, SERPINB13, PMP22, CSTA; and
  - (i) CYBRD1, ARTN, RGS4, CITED2, SLC7A5, ITGB4, CHN1, PI3, TMEM40, NDRG1; and
  - (ii) CYBRD1, ARTN, RGS4, CITED2, SLC7A5; and

(ii) said second set of genes comprises or consists of:
AXL, NOTCH3, GPR30, PROCR, PAX2, PI3KA, CD44, CD133, ALDH1, BMI1, KRT19, NRP1, MSI1, MTOR, JAG1, p38, ESA1, ZEB1, TWIST1, FOXC2, CDH2, ZEB2, FOXC1, SNAI1, SNAI2, TWIST2, GSC, TCF3, CD44, VIM, FN1, MLPH1, ARHGEF11; and
(iii) said third set of genes comprises or consists of:
At least one gene from said first set of genes and at least one of:
AXL, NOTCH3, GPR30, PROCR, PAX2, PI3KA, CD44, CD133, ALDH1, BMI1, KRT19, NRP1, MSI1, MTOR, JAG1, p38, ESA1, ZEB1, TWIST1, FOXC2, CDH2, ZEB2, FOXC1, SNAI1, SNAI2, TWIST2, GSC, TCF3, CD44, VIM, FN1, MLPH1, ARHGEF11; and
(iv) said fourth set of genes comprises or consists of:
At least two genes from said first set of genes and at least one of: AXL, NOTCH3, GPR30, PROCR, PAX2, PI3KA, CD44, CD133, ALDH1, BMI1, KRT19, NRP1, MSI1, MTOR, JAG1, p38, ESA1, ZEB1, TWIST1, FOXC2, CDH2, ZEB2, FOXC1, SNAI1, SNAI2, TWIST2, GSC, TCF3, CD44, VIM, FN1, MLPH1, and ARHGEF11; and
(v) said fifth set of genes comprises or consists of:
beta catenin, FZD1, FZD2, FZD3, FZD4, FZD5, FZD6, FZD7, FZD8, FZD9, FZD10, LRP5, LRP6, WNT1, WNT2, WNT2B, WNT3, WNT3A, WNT4 WNT5A, WNT5B, WNT6, WNT7A, WNT7B, WNT8A, WNT8B, WNT9A, WNT9B, WNT10A, WNT10B, WNT11, WNT16, SRFP1, SRFP2, FRZB, SFRP4, SFRP5, DKK1, DKK2, DKK3, DKK4, WIF1, AKT2, PI3KA, Disheveled, Smoothened, Axin1, Axin2; and
(vi) said sixth set of genes comprises or consists of:
At least one gene from said first set of genes and at least one of:
beta catenin, FZD1, FZD2, FZD3, FZD4, FZD5, FZD6, FZD7, FZD8, FZD9, FZD10, LRP5, LRP6, WNT1, WNT2, WNT2B, WNT3, WNT3A, WNT4 WNT5A, WNT5B, WNT6, WNT7A, WNT7B, WNT8A, WNT8B, WNT9A, WNT9B, WNT10A, WNT10B, WNT11, WNT16, SRFP1, SRFP2, FRZB, SFRP4, SFRP5, DKK1, DKK2, DKK3, DKK4, WIF1, AKT2, PI3KA, Disheveled, Smoothened, Axin1, Axin2; and
(vii) said seventh set of genes comprises or consists of:
TGFbeta1, TGFbeta2, TGFbeta3, ALK1, ALK2, ALK3, ALK4, ALK5, ALK6, ALK7, TGFbetaR-II, BMPR-II, ACTR-IIA, ACTR-IIB, AMHR-II, SMAD2, SMAD3; and
(viii) said eighth set of genes comprises or consists of:
At least one gene from said first set of genes and at least one of:
TGFbeta1, TGFbeta2, TGFbeta3, ALK1, ALK2, ALK3, ALK4, ALK5, ALK6, ALK7, TGFbetaR-II, BMPR-II, ACTR-IIA, ACTR-IIB, AMHR-II, SMAD2, SMAD3; and
(ix) said ninth set of genes comprises or consists of:
S100A4, SDC2, CYP1B1, FERMT2, CYBRD1, SNAI2, CD44, BIN1, PMP22, CTGF, IGFBP3, ANXA6, MAP1B, TAGLN, ENPP2, FBLN5, PCOLCE, FN1, VIM, PPAP2B, SEM5A, VCAN, NID2, NID1, HAS2, PRRX1, TNFAIP6, CLDN4, FADS2, DDIT4, GNG11, PER1, ISCU, KPNA3, SLC44A1, FKBP5, NR2F1, MLPH, DOCK10, RGS4, IGFBP4; and
(x) said tenth set of genes comprises or consists of:
S100A4, SDC2, CYP1B1, FERMT2, CYBRD1, SNAI2, CD44, BIN1, ENPP2, FBLN5, PCOLCE, FN1, VIM, PPAP2B, SEM5A, VCAN, CLDN4, FADS2, DDIT4, GNG11, PER1, ISCU, KPNA3, SLC44A1, FKBP5, IGFBP4; and
(xi) said eleventh set of genes comprises or consists of:
SDC2, CYBRD1, SNAI2, PMP22, CTGF, IGFBP3, ANXA6, MAP1B, TAGLN, PPAP2B, HAS2, NRF2, MLPH, DOCK10, RGS4; and
(xii) said twelfth set of genes comprises or consists of:
FERMT2, BIN1, PMP22, CHN1, CTGF, IGFBP3, NID2, CLDN4, CYBRD1, PRR16, ANXA6, PCOLCE, PPAP2B, HAS2, CDH11, MLPH, DOCK10, RGS4, PVRL3, NID2, SLC44A1, FKBP5; and
(xiii) said thirteenth set of genes comprises or consists of:
FERMT2, BIN1, PMP22, CHN1, CTGF, IGFBP3, NID2, CLDN4; and
(xiv) said fourteenth set of genes comprises or consists of:
HAS2, BIN1, PCOLCE, FERMT2, CTGF, IGFBP3, NID2, SLC44A1, FKBP5, and MLPH; comprising a first agent capable of interacting with a gene expression product of a plurality of genes selected from a first and/or second and/or third and/or fourth and/or fifth and/or sixth set of genes; and wherein the method comprises assaying the level of gene expression product of the plurality of genes.
In one embodiment, the method comprises a second agent capable of interacting with a gene expression product of a plurality of genes in said first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or seventh and/or eighth and/or ninth and/or tenth and/or eleventh and/or twelfth and/or thirteenth and/or fourteenth set of genes. In one embodiment, the method comprises a plurality of agents capable of interacting with a gene expression product of a plurality of genes from said first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or seventh and/or eighth and/or ninth and/or tenth and/or eleventh and/or twelfth and/or thirteenth and/or fourteenth set of genes. In one embodiment, the method comprises a plurality of agents capable of interacting with a plurality of gene expression products of a plurality of genes from said first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or seventh and/or eighth and/or ninth and/or tenth and/or eleventh and/or twelfth and/or thirteenth and/or fourteenth set of genes.
In one embodiment, said agent is a plurality of antibodies. In one embodiment, said agent is a plurality of oligonucleotides. In one embodiment, said gene expression product is a RNA product. In one embodiment, said gene expression product is a protein product. In one embodiment, said agent is a plurality of antibodies. In one embodiment, said agent is a plurality of oligonucleotides. In one embodiment, said gene expression product is a RNA product. In one embodiment, said gene expression product is a protein product. In one embodiment, a value for the level of gene expression product for each gene isoform is assayed. In one embodiment, a value for the level of gene expression product for each gene isoform is assayed by detecting a protein product. In one embodiment, the protein product is detected by an immunoassay, e.g., immunohistochemistry. In one embodiment, a value for the level of gene expression product for each gene isoform is assayed by detecting a RNA product. In one embodiment, the RNA product is detected by a hybridization based method. In one embodiment, the RNA product is detected by microarray. In one embodiment, said microarray is an exon microarray. In one embodiment, the RNA product is detected by a polymerase chain reaction based method. In one embodiment, the RNA product is detected by a sequencing based method. In one embodiment, the RNA product is detected by a quantitative RNA sequencing.
In one embodiment, the gene expression products are derived from a tumor sample, e.g., a preparation of a primary tumor, metastatic tumor, lymph node, circulating tumor cells, ascites, or pleural effusion, plasma, serum, circulating, and interstitial fluid levels.
In one embodiment, a value for the level of gene expression product for each gene is determined. In one embodiment, a value that is a function of the level of gene expression for each gene is determined. In one embodiment, the value is compared to a reference standard, e.g., the level of expression of a control gene in the tumor sample.
In one embodiment, the method further comprises the performance of an algorithm on a computer system to determine a value or values that is a function of a location of a gene expression product in the subject sample and/or a function of a level of a gene expression product of a gene in the subject sample. In one embodiment, the algorithm compares a ratio of the level of gene expression product of at least one of the genes selected from the group: HAS2, BIN1, PCOLCE, FERMT2, CTGF, IGFBP3, NID2, SLC44A1, FKBP5, and MLPH; to the level of gene expression product of at least one of the genes selected from the group: CDH1, and Cytokeratin.
In one embodiment, the method further comprises a plurality of agents capable of interacting with at least one gene expression product selected from the group: CTGF, IGFBP3, TNFAIP6, NID2, HAS2, CCL2, MLPH, NID1, IGFBP4, FBLN5, and PCOLCE. In one embodiment, the method further comprises a plurality of agents capable of interacting with a gene expression product of each gene from the set of genes consisting of: CTGF, IGFBP3, TNFAIP6, NID2, HAS2, CCL2, MLPH, NID1, IGFBP4, FBLN5, and PCOLCE.
In one aspect, methods described herein include methods of assaying in a subject sample the level of gene expression product of a plurality of genes from a first and/or second and/or third and/or fourth and/or fifth and/or sixth set of genes, wherein:

- (i) said first set of genes comprises or consists of:
- S100A4, SDC2, CYP1B1, FERMT2, CYBRD1, SNAI2, CD44, BIN1, PMP22, CTGF, IGFBP3, ANXA6, MAP1B, TAGLN, ENPP2, FBLN5, PCOLCE, FN1, VIM, PPAP2B, SEM5A, VCAN, NID2, NID1, HAS2, PRRX1, TNFAIP6, CLDN4, FADS2, DDIT4, GNG11, PER1, ISCU, KPNA3, SLC44A1, FKBP5, NR2F1, MLPH, DOCK10, RGS4, IGFBP4; and
- (ii) said second set of genes comprises or consists of:
- S100A4, SDC2, CYP1B1, FERMT2, CYBRD1, SNAI2, CD44, BIN1, ENPP2, FBLN5, PCOLCE, FN1, VIM, PPAP2B, SEM5A, VCAN, CLDN4, FADS2, DDIT4, GNG11, PER1, ISCU, KPNA3, SLC44A1, FKBP5, IGFBP4; and
- (iii) said third set of genes comprises or consists of:
- SDC2, CYBRD1, SNAI2, PMP22, CTGF, IGFBP3, ANXA6, MAP1B, TAGLN, PPAP2B, HAS2, NRF2, MLPH, DOCK10, RGS4; and
- (iv) said fourth set of genes comprises or consists of:
- FERMT2, BIN1, PMP22, CHN1, CTGF, IGFBP3, NID2, CLDN4, CYBRD1, PRR16, ANXA6, PCOLCE, PPAP2B, HAS2, CDH11, MLPH, DOCK10, RGS4, PVRL3, NID2, SLC44A1, FKBP5; and
- (v) said fifth set of genes comprises or consists of:
- FERMT2, BIN1, PMP22, CHN1, CTGF, IGFBP3, NID2, CLDN4; and
- (vi) said sixth set of genes comprises or consists of:
- HAS2, BIN1, PCOLCE, FERMT2, CTGF, IGFBP3, NID2, SLC44A1, FKBP5, and MLPH; comprising a first agent capable of interacting with a gene expression product of a plurality of genes selected from a first and/or second and/or third and/or fourth and/or fifth and/or sixth set of genes; and wherein the method comprises assaying the level of gene expression product of the plurality of genes.

In one embodiment, the method comprises a second agent capable of interacting with a gene expression product from said first and/or second and/or third and/or fourth and/or fifth and/or sixth set of genes. In one embodiment, the method comprises a plurality of agents capable of interacting with a gene expression product from said first and/or second and/or third and/or fourth and/or fifth and/or sixth set of genes. In one embodiment, the method comprises a plurality of agents capable of interacting with a plurality of gene expression products from said first and/or second and/or third and/or fourth and/or fifth and/or sixth set of genes.
In one embodiment, said agent is a plurality of antibodies. In one embodiment, said agent is a plurality of oligonucleotides. In one embodiment, said gene expression product is a RNA product. In one embodiment, said gene expression product is a protein product. In one embodiment, the protein product is detected by an immunoassay, e.g., immunohistochemistry. In one embodiment, a value for the level of gene expression product for each gene isoform is assayed by detecting a RNA product. In one embodiment, the RNA product is detected by a hybridization based method. In one embodiment, the RNA product is detected by microarray. In one embodiment, said microarray is an exon microarray. In one embodiment, the RNA product is detected by a polymerase chain reaction based method. In one embodiment, the RNA product is detected by a sequencing based method. In one embodiment, the RNA product is detected by a quantitative RNA sequencing.
In one embodiment, the gene expression products are derived from a tumor sample, e.g., a preparation of a primary tumor, metastatic tumor, lymph node, circulating tumor cells, ascites, or pleural effusion, plasma, serum, circulating, and interstitial fluid levels.
In one embodiment, a value for the level of gene expression product for each gene is determined. In one embodiment, a value that is a function of the level of gene expression for each gene is determined. In one embodiment, the value is compared to a reference standard, e.g., the level of expression of a control gene in the tumor sample.
In one embodiment, the method further comprises the performance of an algorithm on a computer system to determine a value or values that is a function of a location of a gene expression product in the subject sample and/or a function of a level of a gene expression product of a gene in the subject sample. In one embodiment, the algorithm compares a ratio of the level of gene expression product of at least one of the genes selected from the group: HAS2, BIN1, PCOLCE, FERMT2, CTGF, IGFBP3, NID2, SLC44A1, FKBP5, and MLPH; to the level of gene expression product of at least one of the genes selected from the group: CDH1, and Cytokeratin.
In one embodiment, the method further comprises a plurality of agents capable of interacting with at least one gene expression product selected from the group: CTGF, IGFBP3, TNFAIP6, NID2, HAS2, CCL2, MLPH, NID1, IGFBP4, FBLN5, and PCOLCE. In one embodiment, the method further comprises a plurality of agents capable of interacting with a gene expression product of each gene from the set of genes consisting of: CTGF, IGFBP3, TNFAIP6, NID2, HAS2, CCL2, MLPH, NID1, IGFBP4, FBLN5, and PCOLCE.
Reaction Mixtures
In one aspect, reaction mixtures described herein include a reaction mixture comprising: a plurality of detection reagents; and a plurality of target nucleic acid molecules derived from a subject, wherein each of the plurality of detection reagents comprises a plurality probes to measure the level of gene expression product of a gene from a first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or seventh and/or eighth and/or ninth and/or tenth and/or eleventh and/or twelfth and/or thirteenth and/or fourteenth set of genes, wherein:
(i) said first set of genes comprises or consists of:

- KLF13, ANTXR2, IFNAR1, LIX1L, CHST11, AKAP2, PALM2-AKAPT, LOC80298, C7orf2, NUCKS, DKFZP566D1346, LOC388279, FLJ31795, 6orf107, FLJ12439, ENPP2, CITED2, ZEB1, NID2, SEMA5A, DAB2, KCNMA1, PTX3, PCDH9, CREB3L1, BGN, ROR1, DLC1, GNG11, CDH11, NR2F1, PRR16, MYL9, DOCK10, LRIG1, IER3, EML1, NEBL, RGL1, MLPH, DNAJB4, FBLN5, RGS4, HAS2, ITGBL1, IGFBP4, DPT, PCOLCE, GREM1, PPAP2B, CDH2, PMP22, LUM, CHN1, CYP1B1, MME, WNT5A, POSTN, MMP2, CTGF, FBLN1, IGFBP3, DCN, PRRX1, ANXA6, PVRL3, MAP1B, TNFAIP6, CYBRD1, FBN1, NID1, OLFML3, SNAI1, SNAI2, TAGLN, SYNC, TUBA1A, PPM1D, TWIST1, FN1, TGFBR3, SERPINF1, UGDH, SRGN, FAP, PTGER4, PRKCA, FSTL1, MMP1, NRP1, FILIP1L, SCCPDH, LTBP2, XYLT1, HS3ST2, SYT11, TSHZ1, THY1, Sep9, S100A4, TNS3, ENOX1, TGFB1I1, ZEB2, LMCD1, PDGFC, ECM1, TFPI, TBX3, DDR2, PTGER2, CDKN2C, VCAN, CD44, STARD13, SNED1, ZBTB38, SDC2, TPM1, COPZ2, STC1, CDH1, KRT5, KRT6B, CD24, EPCAM, ESRP1, CLDN3, CLDN4, ERBB3, SPOCK1, FERMT2, GLYR1, LTBP1, FADS2, KANK2, PTGFR, COL11A2, KLK3, EIF2C2, ZFP41, FAM49B, PSORS1C2, MRPL42, MRPL54, MRPL47, MRPS23, EIF3S9, ALG5, DNAJC19, TPRXL, NOTCH2, RBM15, ST3 GAL3, NFYA, PCNX, FBXO21, WWOX, CAMK2B, PNPLA2, CLIC5, UGCGL1, FBXL18, ADRBK1, SLC38A2, IL8RA, TAS2R14, CD300LB, GIPC3, RAD51L1, EPPK1, COL1A1, MMP9, SERPINE1, SPARC, TGFB1, TGFB3, TGFBI, TGFBR1, TGIF, TGIF2, THBS1, ANXA5, ACTG1, ARF3, ATP1B3, BAT3, CALD1, CENTD2, CLIC1, CTBS, DPYSL3, DVL3, EXT1, FGFR1, FTL, GNB2L1, GPRC5A, H2AFZ, HIF1A, IL13RA1, KDELR2, LARP1, LPIN2, MARS, MMP10, MMP14, MT2A, MYO10, NUP62, PFKFB3, PLOD2, PSMB7, PSMD8, RIN2, RYBP, SDF4, SETD5, SPP1, STAU1, TUBB3, UBE2S, XPNPEP1, CDKN1A, CHRD, H19, ID3, ID4, IGFBP7, LRP1, MSX1, NOTCH3, PROCR, GBX2, KI67, CCNB1, BUB1, KNTC2, USP22, HCFC1, RNF2, ANK3, FGFR2, CES1, COL1A2, COL3A1, COL5A2, COL6A1, ANKRD25, C10ORF56, C5ORF13, KRT81, NPAC, PLEKHC1, SEPT9, SYNC1, MBP, ABLIM1, ALDH1A3, ALOX15B, ANXA3, AP1M2, ARTN, CA2, CA9, CDH3, CDS1, COL17A1, CORO1A, CTSL2, CXADR, CYP27B1, DSC2, DSG3, DST, EPB41L4B, FGFBP1, FGFR3, FST, GJB3, GRHL2, HBEGF, HOOK1, IL18, IL1B, IRF6, ITGB4, JAG2, KLK10, KLK5, KLK7, KLK8, KRT15, KRT16, KRT17, LEPREL1, MYO5C, NDRG1, NMU, PI3, RAB25, RLN2, RNF128, S100A14, S100A7, S100A8, SERPINB1, SERPINB2, SLC2A9, SLPI, SNX10, SPINT2, SPRR1A, SPRR1B, ST14, SYK, TMEM40, TRIM29, TSPAN1, UCHL1, VSNL1, ZBED2, AQP3, CBARA1, CLCA2, CNTNAP2, CSTA, DSC3, ESRP2, FXYD3, GJB5, GPR87, IL1A, LAMA3, LAMB3, MAP7, MAPK13, MPZL2, NRG1, SCNN1A, SERPINB13, SLC7A5, SPINT1, ST6GALNAC2, TGFA, TP63, TPD52L1, VGLL1, SLC6A8, ABCA12, ANXA8, ANXA8L1, ARHGAP25, ARHGAP8, BDKRB2, BIK, C10ORF10, C10ORF116, C1ORF116, C20ORF19, C6ORF105, CELSR2, CKMT1A, CKMT1B, CLDN1, CST6, CYP4F11, E2F5, ELMO3, EPHA1, EVA1, EXPH5, F11R, FAT2, FLJ12684, FLJ20366, FZD3, GLS2, GNAL, IFI30, IGFBP2, IL1RN, IL4R, IRX4, JUP, KCNK1, KIAA0040, KIAA0888, KIAA1815, KRT14, KRT18, LAD1, LGALS7, LOC553158, LOC653562, LOC728113, LOC729884, LRRC1, LSR, MST1R, MYO1D, NAIP, OCLN, NEFM, NUP62CL, PERP, PLS1, POLR3G, PRKCH, PRRG4, PRSS8, PTPN3, RBM35A, RBM35B, RHBDF2, RHOD, RPS6KA1, RTEL1, SAA1, SAA1, SAA2, SH2D3A, SLC6A10P, SMPDL3B, SNCA, STAC, STAP2, TACSTD1, THBD, TMEM30B, TMPRSS11E, TMPRSS4, TNFRSF6B, TP73L, WWC1, XDH, ZNF165, AXL, GPR30, PAX2, PI3KA, CD133, ALDH1, BMI1, KRT19, MSI1, JAG1, p38, ESA1, FOXC2, FOXC1, TWIST2, GSC, TCF3, MLPH1, ARHGEF11a, DPF2, CASP8, BCL2, SCGN, SWAP70, KIAA0276, C10orf9, C10orf7, ALKBH, TOB2, XPR1, CD59, LRP2, PLP2, MAPK14, CXCL2, MMP7, MGP, MLF1, FLNB, SCNM1, HSPC163, C5orf18, MGC4399, CDW92, TMC4, ZDHHC2, TICAM2, KDELR3, GNPDA1, THEM2, DBR1, FLJ90709, FLJ10774, C16orf33, GAPD, LDHA, MR-1, LARS, GTPBP1, PRSS16, WFDC2, AIM1, DHRS6, DHRS4, GC15429, MGC45840, ECHDC2, GOLGIN-67, AFURS1, KIAA0436, CYP4V2, JTV1, ICMT, DNMT3A, HNMT, METTL7A, METTL2, VIL2, TPD52, ARPC5, NOL8, NSF, RAD23B, SRP54, HSPA2, PBP, THAP2, CIRBP, SNRPN, KIAA0052, DUSP10, SSR1, ERBB4, EMP1, CHPT1, LRPAP1, FLJ11752, CSTF1, KLHL20, DNAJC13, APLP2, ARGBP2, DNAJB1, NEBL, SH3BGRL, NUDT5, GABARAPL1, MAPT, DCBLD1, STK39, PAK2, CSNK2A1, PILRB, ERN1, SGKL, WEE1, MAST4, C11orf17, NUP37, CLTC, COPB2, SLC25A25, ECOP, PDE8A, STAM, TUBB, SNX6, RAB23, PLAA, STC2, LTF, ISGF3G, ATXN3, GTF3C3, GSK3B, KLF10, ELL2, ZBTB20, IRX3, ETS1, SERTAD1, MGC4251, MAFF, SFPQ, CITED4, CEBPD, EIF4E2, HS2ST1, AGPS, PGK1, ATIC, ETNK1, LG2, NCE2, MARCH8, CNOT4, RNF8, PSMA5, DPF2, AMMECR1, KIAA1287, LOC144233, LOC286505, PNAS-4, FLJ20530, HUMPD3, GC45564, CAP350, ETAA16, HAN11, DNAPTP6, C7orf25, FLJ37953, FLJ10587, C7orf36, ELP4, NDEL1, NPD014, KFZP564D172, FAM53C, IER5, LOC255783, KIAA0146, KIAA0792, LOC439994, LOC283481, CG018, LOC130576, NGFRAP1L1, KIAA1217, 4orf7, C21orf86, C9orf64, FLJ13456, FLJ12806, FLJ39370, GATS, CCDC92, FMNL2, ARID1B, ZFHX1B, SSBP2, ARID5B, LOC157381, KPNA3, ARHGAP24, CCND2, VIM, MYCBP2, GAS7, TRAM2, BASP1, FOXO1A, POLR2A, PER1, DDIT4, CD97, BIN1, SH2B3, DDB2, EMP3, NDST2, CHST2, NT5E, PDE4A, CPS1, PTGS1, GGCX, IRF5, ZBTB16, MAP4K4, CHST7, KLF12, NFRKB, PSD, FKSG49, NIFUN, FYN, ZMYM2, CACNA1G, SLC25A16, FLII, EIF1, SEPT6, NPAC, PHF15, NUP188, ABR, CNR1, LOC283824, FSTL4, DNM1, APOBEC3G, ATP2B1, SMPD1, SLC11A1, FXYD5, C14orf139, SH3BGRL3, LUZP1, ZNF335, SH3 KBP1, MST150, PRO1073, LOC388397, FKBP5, HIPK2, DTX1, ST3GAL2, ADAMTS7, TNRC6B, CYGB, SDHAL1, LOC572558, TRIO, FRAS1, KIAA1632, POLS, EBF, MAML2, PTPRA, PLEKHG2, DYM, SOX6, ARHGEF2, ZCCHC6, PPP3CA, FAM70B, TMED5, FLJ43663, HPS1, MEF2A, ST3GAL5, SMYD3, KLF7, LOC200230, RERE, QKI, BICD1, CTNNB1, POU2F2, EIF4ENIF1, BTG1, TCHP, FLJ90709, PCTK2, KIAA1600, B7-H4, PDE4DIP, KIAA0194, HOM-TES; and
- (a) BGN, BIN1, CDH11, CDH2, CHN1, COL1A2, COL3A1, COL5A2, CREB3L1, CTGF, CYBRD1, DCN, DLC1, DNAJB4, DPT, ENPP2, FBLN1, FBLN5, FBN1, GREM1, HAS2, IGFBP3, IGFBP4, MLPH, MME, MMP2, MYL9, NEBL, NID1, NR2F1, OLFML3, PCOLCE, PMP22, POSTN, PPAP2B, PRR16, PRRX1, PTX3, PVRL3, RGL1, RGS4, ROR1, SEMA5A, TNFAIP6, TRAM2, WNT5A, ZEB1, ANXA6, CITED2, COL1A1, COL6A1, CYP1B1, DAB2, DOCK10, EML1, GNG11, IER3, ITGBL1, KCNMA1, LRIG1, LUM, MAP1B, NID2, PCDH9, PRKCA, PTGER4, PTGFR, SDC2, SERPINF1, SNED1, SYT11, TFPI, TGFBR3, VCAN, ANKRD25, C10ORF56, C5ORF13, CCDC92, CDKN2C, COPZ2, DDR2, ECM1, ENOX1, FADS2, FAP, FILIP1L, FSTL1, HS3ST2, KRT81, LMCD1, LTBP1, LTBP2, MMP1, NPAC, NRP1, PDGFC, PLEKHC1, PPM1D, PTGER2, SCCPDH, SEPT9, SPOCK1, SRGN, STARD13, STC1, SYNC1, TAGLN, TBX3, TGFB1I1, THY1, TNS3, TPM1, TSHZ1, TUBA1A, UGDH, VIM, XYLT1, ZBTB38, MBP, ABLIM1, ALDH1A3, ALOX15B, ANXA3, AP1M2, ARTN, CA2, CA9, CAMK2B, CCND2, CDH1, CDH3, CDS1, COL17A1, CORO1A, CTSL2, CXADR, CYP27B1, DSC2, DSG3, DST, EPB41L4B, FGFBP1, FGFR3, FST, GJB3, GRHL2, HBEGF, HOOK1, IL18, IL1B, IRF6, ITGB4, JAG2, KLK10, KLK5, KLK7, KLK8, KRT15, KRT16, KRT17, KRT5, KRT6B, LEPREL1, MYO5C, NDRG1, NMU, PI3, RAB25, RLN2, RNF128, S100A14, S100A7, S100A8, SERPINB1, SERPINB2, SLC2A9, SLPI, SNX10, SPINT2, SPRR1A, SPRR1B, ST14, SYK, TMEM40, TRIM29, TSPAN1, UCHL1, VSNL1, ZBED2, AQP3, CBARA1, CD24, CLCA2, CNTNAP2, CSTA, DSC3, EPCAM, ESRP2, FXYD3, GJB5, GPR87, IL1A, LAMA3, LAMB3, MAP7, MAPK13, MMP9, MPZL2, NRG1, SCNN1A, SERPINB13, SLC7A5, SPINT1, ST6GALNAC2, TGFA, TP63, TPD52L1, VGLL1, SLC6A8, ABCA12, ANK3, ANXA8, ANXA8L1, ARHGAP25, ARHGAP8, BDKRB2, BIK, C10ORF10, C10ORF116, C1ORF116, C20ORF19, C6ORF105, CELSR2, CKMT1A, CKMT1B, CLDN1, CST6, CYP4F11, E2F5, ELMO3, EPHA1, EVA1, EXPH5, F11R, FAT2, FGFR2, FLJ12684, FLJ20366, FZD3, GLS2, GNAL, IFI30, IGFBP2, IL1RN, IL4R, IRX4, JUP, KCNK1, KIAA0040, KIAA0888, KIAA1815, KRT14, KRT18, LAD1, LGALS7, LOC553158, LOC653562, LOC728113, LOC729884, LRRC1, LSR, MST1R, MYO1D, NAIP, OCLN, NEFM, NUP62CL, PERP, PLS1, POLR3G, PRKCH, PRRG4, PRSS8, PTPN3, RBM35A, RBM35B, RHBDF2, RHOD, RPS6KA1, RTEL1, SAA1, SAA1, SAA2, SH2D3A, SLC6A10P, SMPDL3B, SNCA, STAC, STAP2, TACSTD1, THBD, TMEM30B, TMPRSS11E, TMPRSS4, TNFRSF6B, TP73L, WWC1, XDH, ZNF165; and
- (b) BGN, BIN1, CDH11, CDH2, CHN1, COL1A2, COL3A1, COL5A2, CREB3L1, CTGF, CYBRD1, DCN, DLC1, DNAJB4, DPT, ENPP2, FBLN1, FBLN5, FBN1, GREM1, HAS2, IGFBP3, IGFBP4, MLPH, MME, MMP2, MYL9, NEBL, NID1, NR2F1, OLFML3, PCOLCE, PMP22, POSTN, PPAP2B, PRR16, PRRX1, PTX3, PVRL3, RGL1, RGS4, ROR1, SEMA5A, TNFAIP6, TRAM2, WNT5A, ZEB1, ANXA6, CITED2, COL1A1, COL6A1, CYP1B1, DAB2, DOCK10, EML1, GNG11, IER3, ITGBL1, KCNMA1, LRIG1, LUM, MAP1B, NID2, PCDH9, PRKCA, PTGER4, PTGFR, SDC2, SERPINF1, SNED1, SYT11, TFPI, TGFBR3, VCAN, ANKRD25, C10ORF56, C5ORF13, CCDC92, CDKN2C, COPZ2, DDR2, ECM1, ENOX1, FADS2, FAP, FILIP1L, FSTL1, HS3ST2, KRT81, LMCD1, LTBP1, LTBP2, MMP1, NPAC, NRP1, PDGFC, PLEKHC1, PPM1D, PTGER2, SCCPDH, SEPT9, SPOCK1, SRGN, STARD13, STC1, SYNC1, TAGLN, TBX3, TGFB1I1, THY1, TNS3, TPM1, TSHZ1, TUBA1A, UGDH, VIM, XYLT1, ZBTB38; and
- (c) ENPP2, CITED2, ZEB1, NID2, SEMA5A, DAB2, KCNMA1, PTX3, PCDH9, CREB3L1, BGN, ROR1, DLC1, GNG11, CDH11, NR2F1, NR2F2, PRR16, MYL9, DOCK10, LRIG1, IER3, EML1, NEBL, RGL1, MLPH, TRAM2, DNAJB4, FBLN5, RGS4, HAS2, ITGBL1, IGFBP4, BIN1, DPT, PCOLCE, GREM1, PPAP2B, CDH2, PMP22, LUM, CHN1, CYP1B1, MME, WNT5A, POSTN, MMP2, CTGF, FBLN1, IGFBP3, DCN, PRRX1, ANXA6, PVRL3, MAP1B, TNFAIP6, CYBRD1, FBN1, NID1, OLFML3, SNAI1, SNAI2, TAGLN, SYNC, TUBA1A, PPM1D, TWIST1, FN1, TGFBR3, SERPINF1, UGDH, SRGN, FAP, PTGER4, VIM, PRKCA, FSTL1, MMP1, NRP1, FILIP1L, SCCPDH, LTBP2, XYLT1, HS3ST2, SYT11, TSHZ1, THY1, SEPT9, S100A4, CCDC92, TNS3, ENOX1, TGFB1I1, ZEB2, LMCD1, PDGFC, ECM1, TFPI, TBX3, DDR2, PTGER2, CDKN2C, VCAN, CD44, STARD13, SNED1, ZBTB38, SDC2, TPM1, COPZ2, STC1, CDH1, KRT5, KRT6B, CD24, EPCAM, ESRP1, CLDN3, CLDN4, ERBB3, SPOCK1, FERMT2, GLYR1, LTBP1, FADS2, KANK2, PTGFR, ARTN, SLC7A5, EPB41L4B, KRT15, CDH3, TPD52L1, ANXA3, PI3, NDRG1, DSG3, S100A14, S100S7, KLK8, S100A8, PTX3, SPINT2, FXYD3, KRT16, RAB25, LGALS7, DSC3, COL5A2, ITGB4, TMEM40, SPRR1A, AP1M2, SERPINB13, TP73L; and
- (d) CYBRD1, ARTN, CITED2, SLC7A5, CHN1, CTGF, CSTA, MYL9, EPB41L4B, KRT15, LRIG1, CDH3, TPD52L1, ANXA3, OLFML3, CORO1A, PI3, NDRG1, DSG3, PMP22, S100A14, S100S7, KLK8, S100A8, PTX3, SPINT2, IER3, FXYD3, KRT16, TGFBR3, KRT5, RAB25, SDC2, LUM, LGALS7, KRT6B, CD24, COL1A2, GNG11, DSC3, COL5A2, RGS4, ITGB4, TMEM40, SPRR1A, AP1M2, SERPINB13, SNED1, LEPREL1, TRAM2, TP73L, and PAP2B; and
- (e) CYBRD1, ARTN, CITED2, SLC7A5, CHN1, CTGF, CSTA, MYL9, EPB41L4B, KRT15, LRIG1, CDH3, TPD52L1, ANXA3, OLFML3, CORO1A; and
- (f) PI3, NDRG1, DSG3, PMP22, S100A14, S100S7, KLK8, S100A8, PTX3, SPINT2, IER3, FXYD3, KRT16, TGFBR3; and
- (g) KRT5, RAB25, SDC2, LUM, LGALS7, KRT6B, CD24, COL1A2, GNG11, DSC3, COL5A2; and
- (h) RGS4, ITGB4, TMEM40, SPRR1A, AP1M2, AERPINB13, SNED1, LEPREL1, TRAM2, TP73L, and PAB2B; and
- (i) CYBRD1, ARTN, KRT15, ITGB4, RGS4, SPRR1B, SLC7A5, ANXA3, IL18, AP1M2, CHN1, OLFML3, ZBED2, LEPREL1, ABLIM1, CDH3, DNAJB4, TRAM2, UCHL1, CTGF; and
  - (i) CYBRD1, ARTN, KRT15, ITGB4, RGS4, SPRR1B, SLC7A5, ANXA3, IL18, AP1M2; and
  - (ii) CYBRD1, ARTN, KRT15, ITGB4, RGS4; and
- (j) CYBRD1, RGS4, CDH3, CHN1, SLC7A5, PRRX1, CITED2, COL5A2, ARTN, ZBED2, SNED1, ITGB4, LUM, PVRL3, OLFML3, BIN1, CCND2, DAB2, ANXA3, IL18; and
  - (i) CYBRD1, RGS4, CDH3, CHN1, SLC7A5, PRRX1, CITED2, COL5A2, ARTN, ZBED2; and
  - (ii) CYBRD1, RGS4, CDH3, CHN1, SLC7A5; and
- (k) CYBRD1, ARTN, RGS4, CITED2, SLC7A5, ITGB4, CHN1, PI3, TMEM40, NDRG1, SPRR1A, PCDH9, CTGF, MAP1B, DSG3, AP1M2, FBLN5, SERPINB13, PMP22, CSTA; and
  - (i) CYBRD1, ARTN, RGS4, CITED2, SLC7A5, ITGB4, CHN1, PI3, TMEM40, NDRG1; and
  - (ii) CYBRD1, ARTN, RGS4, CITED2, SLC7A5; and

(ii) said second set of genes comprises or consists of:
AXL, NOTCH3, GPR30, PROCR, PAX2, PI3KA, CD44, CD133, ALDH1, BMI1, KRT19, NRP1, MSI1, MTOR, JAG1, p38, ESA1, ZEB1, TWIST1, FOXC2, CDH2, ZEB2, FOXC1, SNAI1, SNAI2, TWIST2, GSC, TCF3, CD44, VIM, FN1, MLPH1, ARHGEF11; and
(iii) said third set of genes comprises or consists of:
At least one gene from said first set of genes and at least one of:
AXL, NOTCH3, GPR30, PROCR, PAX2, PI3KA, CD44, CD133, ALDH1, BMI1, KRT19, NRP1, MSI1, MTOR, JAG1, p38, ESA1, ZEB1, TWIST1, FOXC2, CDH2, ZEB2, FOXC1, SNAI1, SNAI2, TWIST2, GSC, TCF3, CD44, VIM, FN1, MLPH1, ARHGEF11; and
(iv) said fourth set of genes comprises or consists of:
At least two genes from said first set of genes and at least one of: AXL, NOTCH3, GPR30, PROCR, PAX2, PI3KA, CD44, CD133, ALDH1, BMI1, KRT19, NRP1, MSI1, MTOR, JAG1, p38, ESA1, ZEB1, TWIST1, FOXC2, CDH2, ZEB2, FOXC1, SNAI1, SNAI2, TWIST2, GSC, TCF3, CD44, VIM, FN1, MLPH1, and ARHGEF11; and
(v) said fifth set of genes comprises or consists of:
beta catenin, FZD1, FZD2, FZD3, FZD4, FZD5, FZD6, FZD7, FZD8, FZD9, FZD10, LRP5, LRP6, WNT1, WNT2, WNT2B, WNT3, WNT3A, WNT4 WNT5A, WNT5B, WNT6, WNT7A, WNT7B, WNT8A, WNT8B, WNT9A, WNT9B, WNT10A, WNT10B, WNT11, WNT16, SRFP1, SRFP2, FRZB, SFRP4, SFRP5, DKK1, DKK2, DKK3, DKK4, WIF1, AKT2, PI3KA, Disheveled, Smoothened, Axin1, Axin2; and
(vi) said sixth set of genes comprises or consists of:
At least one gene from said first set of genes and at least one of:
beta catenin, FZD1, FZD2, FZD3, FZD4, FZD5, FZD6, FZD7, FZD8, FZD9, FZD10, LRP5, LRP6, WNT1, WNT2, WNT2B, WNT3, WNT3A, WNT4 WNT5A, WNT5B, WNT6, WNT7A, WNT7B, WNT8A, WNT8B, WNT9A, WNT9B, WNT10A, WNT10B, WNT11, WNT16, SRFP1, SRFP2, FRZB, SFRP4, SFRP5, DKK1, DKK2, DKK3, DKK4, WIF1, AKT2, PI3KA, Disheveled, Smoothened, Axin1, Axin2; and
(vii) said seventh set of genes comprises or consists of:
TGFbeta1, TGFbeta2, TGFbeta3, ALK1, ALK2, ALK3, ALK4, ALK5, ALK6, ALK7, TGFbetaR-II, BMPR-II, ACTR-IIA, ACTR-IIB, AMHR-II, SMAD2, SMAD3; and
(viii) said eighth set of genes comprises or consists of:
At least one gene from said first set of genes and at least one of:
TGFbeta1, TGFbeta2, TGFbeta3, ALK1, ALK2, ALK3, ALK4, ALK5, ALK6, ALK7, TGFbetaR-II, BMPR-II, ACTR-IIA, ACTR-IIB, AMHR-II, SMAD2, SMAD3; and
(ix) said ninth set of genes comprises or consists of:
S100A4, SDC2, CYP1B1, FERMT2, CYBRD1, SNAI2, CD44, BIN1, PMP22, CTGF, IGFBP3, ANXA6, MAP1B, TAGLN, ENPP2, FBLN5, PCOLCE, FN1, VIM, PPAP2B, SEM5A, VCAN, NID2, NID1, HAS2, PRRX1, TNFAIP6, CLDN4, FADS2, DDIT4, GNG11, PER1, ISCU, KPNA3, SLC44A1, FKBP5, NR2F1, MLPH, DOCK10, RGS4, IGFBP4; and
(x) said tenth set of genes comprises or consists of:
S100A4, SDC2, CYP1B1, FERMT2, CYBRD1, SNAI2, CD44, BIN1, ENPP2, FBLN5, PCOLCE, FN1, VIM, PPAP2B, SEM5A, VCAN, CLDN4, FADS2, DDIT4, GNG11, PER1, ISCU, KPNA3, SLC44A1, FKBP5, IGFBP4; and
(xi) said eleventh set of genes comprises or consists of:
SDC2, CYBRD1, SNAI2, PMP22, CTGF, IGFBP3, ANXA6, MAP1B, TAGLN, PPAP2B, HAS2, NRF2, MLPH, DOCK10, RGS4; and
(xii) said twelfth set of genes comprises or consists of:
FERMT2, BIN1, PMP22, CHN1, CTGF, IGFBP3, NID2, CLDN4, CYBRD1, PRR16, ANXA6, PCOLCE, PPAP2B, HAS2, CDH11, MLPH, DOCK10, RGS4, PVRL3, NID2, SLC44A1, FKBP5; and
(xiii) said thirteenth set of genes comprises or consists of:
FERMT2, BIN1, PMP22, CHN1, CTGF, IGFBP3, NID2, CLDN4; and
(xiv) said fourteenth set of genes comprises or consists of:
HAS2, BIN1, PCOLCE, FERMT2, CTGF, IGFBP3, NID2, SLC44A1, FKBP5, and MLPH.
In one embodiment, each probe comprises a DNA, RNA or mixed DNA/RNA molecule, which is complementary to a nucleic acid sequence on each of the plurality of target nucleic acid molecules, wherein each target nucleic acid molecule is derived from a gene in said first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or seventh and/or eighth and/or ninth and/or tenth and/or eleventh and/or twelfth and/or thirteenth and/or fourteenth set of genes. In one embodiment, each of the plurality of detection reagents comprises a probe to measure the expression at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 genes in said first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or seventh and/or eighth and/or ninth and/or tenth and/or eleventh and/or twelfth and/or thirteenth and/or fourteenth set of genes. In one embodiment, each of the plurality of detection reagents comprises a probe to measure the expression 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or genes in said first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or seventh and/or eighth and/or ninth and/or tenth and/or eleventh and/or twelfth and/or thirteenth and/or fourteenth set of genes. In one embodiment, each of the plurality of detection reagents comprises a probe to measure the expression of no more than 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 genes in said first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or seventh and/or eighth and/or ninth and/or tenth and/or eleventh and/or twelfth and/or thirteenth and/or fourteenth set of genes. In one embodiment, each of the plurality of detection reagents comprises a probe to measure the expression of only genes in said first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or seventh and/or eighth and/or ninth and/or tenth and/or eleventh and/or twelfth and/or thirteenth and/or fourteenth set of genes.
In an embodiment, the probe is a nucleic acid molecule. In one embodiment, the plurality of target nucleic acid molecules is derived from a subject with cancer. Also described herein are kits comprising detection reagents described herein.
In one aspect, reaction mixtures described herein include a reaction mixture comprising:
a plurality of detection reagents, e.g., a plurality of substrates, e.g., a plurality of antibodies; and a plurality of target proteins derived from a cancer, wherein each of the plurality of target proteins is encoded by a gene in said first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or seventh and/or eighth and/or ninth and/or tenth and/or eleventh and/or twelfth and/or thirteenth and/or fourteenth set of genes, and wherein each of the plurality of detection reagents is a probe specific for one of the plurality of target proteins, e.g., binds to the target protein.
In one embodiment, each of the plurality of detection reagents comprises a probe to measure the expression at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 genes in said first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or seventh and/or eighth and/or ninth and/or tenth and/or eleventh and/or twelfth and/or thirteenth and/or fourteenth set of genes. In one embodiment, each of the plurality of detection reagents comprises a probe to measure the expression 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 genes in said first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or seventh and/or eighth and/or ninth and/or tenth and/or eleventh and/or twelfth and/or thirteenth and/or fourteenth set of genes. In one embodiment, each of the plurality of detection reagents comprises a probe to measure the expression of no more than 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 genes in said first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or seventh and/or eighth and/or ninth and/or tenth and/or eleventh and/or twelfth and/or thirteenth and/or fourteenth set of genes. In one embodiment, each of the plurality of detection reagents comprises a probe to measure the expression of only genes in said first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or seventh and/or eighth and/or ninth and/or tenth and/or eleventh and/or twelfth and/or thirteenth and/or fourteenth set of genes.
In one embodiment, the plurality of target proteins is derived from a patient with a cancer. Also described herein are kits comprising detection reagents described herein.
Also described herein are methods of making a reaction mixture.
In one aspect, described herein are methods of making a reaction mixture comprising:
combining a plurality of detection reagents, with a plurality of target nucleic acid molecules derived from a patient with an ovarian cancer, wherein each target nucleic acid molecule is derived from a plurality of genes in Table 1, and wherein each of the plurality of detection reagents comprises a probe to measure the expression of a gene in said first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or seventh and/or eighth and/or ninth and/or tenth and/or eleventh and/or twelfth and/or thirteenth and/or fourteenth set of genes.
In one aspect, described herein are methods of making a reaction mixture comprising:
combining a plurality of detection reagents, e.g., a plurality of substrates, e.g., a plurality of antibodies; and a plurality of target proteins derived from an ovarian cancer, wherein each of the plurality of target proteins is encoded by a gene in said first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or seventh and/or eighth and/or ninth and/or tenth and/or eleventh and/or twelfth and/or thirteenth and/or fourteenth set of genes, and wherein each of the plurality of detection reagents is a probe specific for one of the plurality of target proteins, e.g., binds to the target protein.
In one aspect, reaction mixtures described herein include a reaction mixture comprising: a plurality of detection reagents; and a plurality of target nucleic acid molecules derived from a subject, wherein each of the plurality of detection reagents comprises a plurality probes to measure the level of gene expression product of a gene from a first and/or second and/or third and/or fourth and/or fifth and/or sixth set of genes, wherein:

- (i) said first set of genes comprises or consists of:
  S100A4, SDC2, CYP1B1, FERMT2, CYBRD1, SNAI2, CD44, BIN1, PMP22, CTGF, IGFBP3, ANXA6, MAP1B, TAGLN, ENPP2, FBLN5, PCOLCE, FN1, VIM, PPAP2B, SEM5A, VCAN, NID2, NID1, HAS2, PRRX1, TNFAIP6, CLDN4, FADS2, DDIT4, GNG11, PER1, ISCU, KPNA3, SLC44A1, FKBP5, NR2F1, MLPH, DOCK10, RGS4, IGFBP4; and
- (ii) said second set of genes comprises or consists of:
  S100A4, SDC2, CYP1B1, FERMT2, CYBRD1, SNAI2, CD44, BIN1, ENPP2, FBLN5, PCOLCE, FN1, VIM, PPAP2B, SEM5A, VCAN, CLDN4, FADS2, DDIT4, GNG11, PER1, ISCU, KPNA3, SLC44A1, FKBP5, IGFBP4; and
- (iii) said third set of genes comprises or consists of:

SDC2, CYBRD1, SNAI2, PMP22, CTGF, IGFBP3, ANXA6, MAP1B, TAGLN, PPAP2B, HAS2, NRF2, MLPH, DOCK10, RGS4; and

- (iv) said fourth set of genes comprises or consists of:

FERMT2, BIN1, PMP22, CHN1, CTGF, IGFBP3, NID2, CLDN4, CYBRD1, PRR16, ANXA6, PCOLCE, PPAP2B, HAS2, CDH11, MLPH, DOCK10, RGS4, PVRL3, NID2, SLC44A1, FKBP5; and

- (v) said fifth set of genes comprises or consists of:

FERMT2, BIN1, PMP22, CHN1, CTGF, IGFBP3, NID2, CLDN4; and

- (vi) said sixth set of genes comprises or consists of:
- HAS2, BIN1, PCOLCE, FERMT2, CTGF, IGFBP3, NID2, SLC44A1, FKBP5, and MLPH.

In one embodiment, each probe comprises a DNA, RNA or mixed DNA/RNA molecule, which is complementary to a nucleic acid sequence on each of the plurality of target nucleic acid molecules, wherein each target nucleic acid molecule is derived from a gene in said first and/or second and/or third and/or fourth and/or fifth and/or sixth set of genes. In one embodiment, each of the plurality of detection reagents comprises a probe to measure the expression at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 genes in said first and/or second and/or third and/or fourth and/or fifth and/or sixth set of genes. In one embodiment, each of the plurality of detection reagents comprises a probe to measure the expression 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 genes in said first and/or second and/or third and/or fourth and/or fifth and/or sixth set of genes. In one embodiment, each of the plurality of detection reagents comprises a probe to measure the expression of no more than 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 genes in said first and/or second and/or third and/or fourth and/or fifth and/or sixth set of genes. In one embodiment, each of the plurality of detection reagents comprises a probe to measure the expression of only genes in said first and/or second and/or third and/or fourth and/or fifth and/or sixth set of genes.
In an embodiment, the probe is a nucleic acid molecule. In one embodiment, the plurality of target nucleic acid molecules is derived from a subject with cancer. Also described herein are kits comprising detection reagents described herein.
In one aspect, reaction mixtures described herein include a reaction mixture comprising:
a plurality of detection reagents, e.g., a plurality of substrates, e.g., a plurality of antibodies; and a plurality of target proteins derived from a cancer, wherein each of the plurality of target proteins is encoded by a gene in a first and/or second and/or third and/or fourth and/or fifth and/or sixth set of genes, wherein:

- (i) said first set of genes comprises or consists of:
- S100A4, SDC2, CYP1B1, FERMT2, CYBRD1, SNAI2, CD44, BIN1, PMP22, CTGF, IGFBP3, ANXA6, MAP1B, TAGLN, ENPP2, FBLN5, PCOLCE, FN1, VIM, PPAP2B, SEM5A, VCAN, NID2, NID1, HAS2, PRRX1, TNFAIP6, CLDN4, FADS2, DDIT4, GNG11, PER1, ISCU, KPNA3, SLC44A1, FKBP5, NR2F1, MLPH, DOCK10, RGS4, IGFBP4; and
- (ii) said second set of genes comprises or consists of:
- S100A4, SDC2, CYP1B1, FERMT2, CYBRD1, SNAI2, CD44, BIN1, ENPP2, FBLN5, PCOLCE, FN1, VIM, PPAP2B, SEM5A, VCAN, CLDN4, FADS2, DDIT4, GNG11, PER1, ISCU, KPNA3, SLC44A1, FKBP5, IGFBP4; and
- (iii) said third set of genes comprises or consists of:
- SDC2, CYBRD1, SNAI2, PMP22, CTGF, IGFBP3, ANXA6, MAP1B, TAGLN, PPAP2B, HAS2, NRF2, MLPH, DOCK10, RGS4; and
- (iv) said fourth set of genes comprises or consists of:
- FERMT2, BIN1, PMP22, CHN1, CTGF, IGFBP3, NID2, CLDN4, CYBRD1, PRR16, ANXA6, PCOLCE, PPAP2B, HAS2, CDH11, MLPH, DOCK10, RGS4, PVRL3, NID2, SLC44A1, FKBP5; and
- (v) said fifth set of genes comprises or consists of:
- FERMT2, BIN1, PMP22, CHN1, CTGF, IGFBP3, NID2, CLDN4; and
- (vi) said sixth set of genes comprises or consists of:

HAS2, BIN1, PCOLCE, FERMT2, CTGF, IGFBP3, NID2, SLC44A1, FKBP5, and MLPH;
and wherein each of the plurality of detection reagents is a probe specific for one of the plurality of target proteins, e.g., binds to the target protein.
In one embodiment, each of the plurality of detection reagents comprises a probe to measure the expression at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 genes in said first and/or second and/or third and/or fourth and/or fifth and/or sixth set of genes. In one embodiment, each of the plurality of detection reagents comprises a probe to measure the expression 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 genes in said first and/or second and/or third and/or fourth and/or fifth and/or sixth set of genes. In one embodiment, each of the plurality of detection reagents comprises a probe to measure the expression of no more than 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 genes in said first and/or second and/or third and/or fourth and/or fifth and/or sixth set of genes. In one embodiment, each of the plurality of detection reagents comprises a probe to measure the expression of only genes in said first and/or second and/or third and/or fourth and/or fifth and/or sixth set of genes.
In one embodiment, the plurality of target proteins is derived from a patient with a cancer. Also described herein are kits comprising detection reagents described herein.
Also described herein are methods of making a reaction mixture.
In one aspect, described herein are methods of making a reaction mixture comprising:
combining a plurality of detection reagents, with a plurality of target nucleic acid molecules derived from a patient with an ovarian cancer, wherein each target nucleic acid molecule is derived from a plurality of genes in a first and/or second and/or third and/or fourth and/or fifth and/or sixth set of genes, wherein:

- (i) said first set of genes comprises or consists of:
- S100A4, SDC2, CYP1B1, FERMT2, CYBRD1, SNAI2, CD44, BIN1, PMP22, CTGF, IGFBP3, ANXA6, MAP1B, TAGLN, ENPP2, FBLN5, PCOLCE, FN1, VIM, PPAP2B, SEM5A, VCAN, NID2, NID1, HAS2, PRRX1, TNFAIP6, CLDN4, FADS2, DDIT4, GNG11, PER1, ISCU, KPNA3, SLC44A1, FKBP5, NR2F1, MLPH, DOCK10, RGS4, IGFBP4; and
- (ii) said second set of genes comprises or consists of:
- S100A4, SDC2, CYP1B1, FERMT2, CYBRD1, SNAI2, CD44, BIN1, ENPP2, FBLN5, PCOLCE, FN1, VIM, PPAP2B, SEM5A, VCAN, CLDN4, FADS2, DDIT4, GNG11, PER1, ISCU, KPNA3, SLC44A1, FKBP5, IGFBP4; and
- (iii) said third set of genes comprises or consists of:
- SDC2, CYBRD1, SNAI2, PMP22, CTGF, IGFBP3, ANXA6, MAP1B, TAGLN, PPAP2B, HAS2, NRF2, MLPH, DOCK10, RGS4; and
- (iv) said fourth set of genes comprises or consists of:
- FERMT2, BIN1, PMP22, CHN1, CTGF, IGFBP3, NID2, CLDN4, CYBRD1, PRR16, ANXA6, PCOLCE, PPAP2B, HAS2, CDH11, MLPH, DOCK10, RGS4, PVRL3, NID2, SLC44A1, FKBP5; and
- (v) said fifth set of genes comprises or consists of:
- FERMT2, BIN1, PMP22, CHN1, CTGF, IGFBP3, NID2, CLDN4; and
- (vi) said sixth set of genes comprises or consists of:
- HAS2, BIN1, PCOLCE, FERMT2, CTGF, IGFBP3, NID2, SLC44A1, FKBP5, and MLPH;

and wherein each of the plurality of detection reagents comprises a probe to measure the expression of a gene in said first and/or second and/or third and/or fourth and/or fifth and/or sixth set of genes.
In one aspect, described herein are methods of making a reaction mixture comprising:
combining a plurality of detection reagents, e.g., a plurality of substrates, e.g., a plurality of antibodies; and a plurality of target proteins derived from an ovarian cancer, wherein each of the plurality of target proteins is encoded by a gene in a first and/or second and/or third and/or fourth and/or fifth and/or sixth set of genes, wherein:

and wherein each of the plurality of detection reagents is a probe specific for one of the plurality of target proteins, e.g., binds to the target protein.
In Vitro Assays
Also described herein are in vitro methods and assays. In one aspect described herein are in vitro methods and assays of determining if a subject is a potential candidate for treatment with an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells, the method comprising determining the level of gene expression product of a plurality of genes selected from a first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or seventh and/or eighth and/or ninth and/or tenth and/or eleventh and/or twelfth and/or thirteenth and/or fourteenth set of genes, wherein:
(i) said first set of genes comprises or consists of:

- KLF13, ANTXR2, IFNAR1, LIX1L, CHST11, AKAP2, PALM2-AKAPT, LOC80298, C7 orf2, NUCKS, DKFZP566D1346, LOC388279, FLJ31795, 6orf107, FLJ12439, ENPP2, CITED2, ZEB1, NID2, SEMA5A, DAB2, KCNMA1, PTX3, PCDH9, CREB3L1, BGN, ROR1, DLC1, GNG11, CDH11, NR2F1, PRR16, MYL9, DOCK10, LRIG1, IER3, EML1, NEBL, RGL1, MLPH, DNAJB4, FBLN5, RGS4, HAS2, ITGBL1, IGFBP4, DPT, PCOLCE, GREM1, PPAP2B, CDH2, PMP22, LUM, CHN1, CYP1B1, MME, WNT5A, POSTN, MMP2, CTGF, FBLN1, IGFBP3, DCN, PRRX1, ANXA6, PVRL3, MAP1B, TNFAIP6, CYBRD1, FBN1, NID1, OLFML3, SNAI1, SNAI2, TAGLN, SYNC, TUBA1A, PPM1D, TWIST1, FN1, TGFBR3, SERPINF1, UGDH, SRGN, FAP, PTGER4, PRKCA, FSTL1, MMP1, NRP1, FILIP1L, SCCPDH, LTBP2, XYLT1, HS3ST2, SYT11, TSHZ1, THY1, Sep9, S100A4, TNS3, ENOX1, TGFB1I1, ZEB2, LMCD1, PDGFC, ECM1, TFPI, TBX3, DDR2, PTGER2, CDKN2C, VCAN, CD44, STARD13, SNED1, ZBTB38, SDC2, TPM1, COPZ2, STC1, CDH1, KRT5, KRT6B, CD24, EPCAM, ESRP1, CLDN3, CLDN4, ERBB3, SPOCK1, FERMT2, GLYR1, LTBP1, FADS2, KANK2, PTGFR, COL11A2, KLK3, EIF2C2, ZFP41, FAM49B, PSORS1C2, MRPL42, MRPL54, MRPL47, MRPS23, EIF3S9, ALG5, DNAJC19, TPRXL, NOTCH2, RBM15, ST3 GAL3, NFYA, PCNX, FBXO21, WWOX, CAMK2B, PNPLA2, CLIC5, UGCGL1, FBXL18, ADRBK1, SLC38A2, IL8RA, TAS2R14, CD300LB, GIPC3, RAD51L1, EPPK1, COL1A1, MMP9, SERPINE1, SPARC, TGFB1, TGFB3, TGFBI, TGFBR1, TGIF, TGIF2, THBS1, ANXA5, ACTG1, ARF3, ATP1B3, BAT3, CALD1, CENTD2, CLIC1, CTBS, DPYSL3, DVL3, EXT1, FGFR1, FTL, GNB2L1, GPRC5A, H2AFZ, HIF1A, IL13RA1, KDELR2, LARP1, LPIN2, MARS, MMP10, MMP14, MT2A, MYO10, NUP62, PFKFB3, PLOD2, PSMB7, PSMD8, RIN2, RYBP, SDF4, SETD5, SPP1, STAU1, TUBB3, UBE2S, XPNPEP1, CDKN1A, CHRD, H19, ID3, ID4, IGFBP7, LRP1, MSX1, NOTCH3, PROCR, GBX2, KI67, CCNB1, BUB1, KNTC2, USP22, HCFC1, RNF2, ANK3, FGFR2, CES1, COL1A2, COL3A1, COL5A2, COL6A1, ANKRD25, C10ORF56, C5ORF13, KRT81, NPAC, PLEKHC1, SEPT9, SYNC1, MBP, ABLIM1, ALDH1A3, ALOX15B, ANXA3, AP1M2, ARTN, CA2, CA9, CDH3, CDS1, COL17A1, CORO1A, CTSL2, CXADR, CYP27B1, DSC2, DSG3, DST, EPB41L4B, FGFBP1, FGFR3, FST, GJB3, GRHL2, HBEGF, HOOK1, IL18, IL1B, IRF6, ITGB4, JAG2, KLK10, KLK5, KLK7, KLK8, KRT15, KRT16, KRT17, LEPREL1, MYO5C, NDRG1, NMU, PI3, RAB25, RLN2, RNF128, S100A14, S100A7, S100A8, SERPINB1, SERPINB2, SLC2A9, SLPI, SNX10, SPINT2, SPRR1A, SPRR1B, ST14, SYK, TMEM40, TRIM29, TSPAN1, UCHL1, VSNL1, ZBED2, AQP3, CBARA1, CLCA2, CNTNAP2, CSTA, DSC3, ESRP2, FXYD3, GJB5, GPR87, IL1A, LAMA3, LAMB3, MAP7, MAPK13, MPZL2, NRG1, SCNN1A, SERPINB13, SLC7A5, SPINT1, ST6GALNAC2, TGFA, TP63, TPD52L1, VGLL1, SLC6A8, ABCA12, ANXA8, ANXA8L1, ARHGAP25, ARHGAP8, BDKRB2, BIK, C10ORF10, C10ORF116, C1ORF116, C20ORF19, C6ORF105, CELSR2, CKMT1A, CKMT1B, CLDN1, CST6, CYP4F11, E2F5, ELMO3, EPHA1, EVA1, EXPH5, F11R, FAT2, FLJ12684, FLJ20366, FZD3, GLS2, GNAL, IFI30, IGFBP2, IL1RN, IL4R, IRX4, JUP, KCNK1, KIAA0040, KIAA0888, KIAA1815, KRT14, KRT18, LAD1, LGALS7, LOC553158, LOC653562, LOC728113, LOC729884, LRRC1, LSR, MST1R, MYO1D, NAIP, OCLN, NEFM, NUP62CL, PERP, PLS1, POLR3G, PRKCH, PRRG4, PRSS8, PTPN3, RBM35A, RBM35B, RHBDF2, RHOD, RPS6KA1, RTEL1, SAA1, SAA1, SAA2, SH2D3A, SLC6A10P, SMPDL3B, SNCA, STAC, STAP2, TACSTD1, THBD, TMEM30B, TMPRSS11E, TMPRSS4, TNFRSF6B, TP73L, WWC1, XDH, ZNF165, AXL, GPR30, PAX2, PI3KA, CD133, ALDH1, BMI1, KRT19, MSI1, JAG1, p38, ESA1, FOXC2, FOXC1, TWIST2, GSC, TCF3, MLPH1, ARHGEF11a, DPF2, CASP8, BCL2, SCGN, SWAP70, KIAA0276, C10orf9, C10orf7, ALKBH, TOB2, XPR1, CD59, LRP2, PLP2, MAPK14, CXCL2, MMP7, MGP, MLF1, FLNB, SCNM1, HSPC163, C5orf18, MGC4399, CDW92, TMC4, ZDHHC2, TICAM2, KDELR3, GNPDA1, THEM2, DBR1, FLJ90709, FLJ10774, C16orf33, GAPD, LDHA, MR-1, LARS, GTPBP1, PRSS16, WFDC2, AIM1, DHRS6, DHRS4, GC15429, MGC45840, ECHDC2, GOLGIN-67, AFURS1, KIAA0436, CYP4V2, JTV1, ICMT, DNMT3A, HNMT, METTL7A, METTL2, VIL2, TPD52, ARPC5, NOL8, NSF, RAD23B, SRP54, HSPA2, PBP, THAP2, CIRBP, SNRPN, KIAA0052, DUSP10, SSR1, ERBB4, EMP1, CHPT1, LRPAP1, FLJ11752, CSTF1, KLHL20, DNAJC13, APLP2, ARGBP2, DNAJB1, NEBL, SH3BGRL, NUDT5, GABARAPL1, MAPT, DCBLD1, STK39, PAK2, CSNK2A1, PILRB, ERN1, SGKL, WEE1, MAST4, C11orf17, NUP37, CLTC, COPB2, SLC25A25, ECOP, PDE8A, STAM, TUBB, SNX6, RAB23, PLAA, STC2, LTF, ISGF3G, ATXN3, GTF3C3, GSK3B, KLF10, ELL2, ZBTB20, IRX3, ETS1, SERTAD1, MGC4251, MAFF, SFPQ, CITED4, CEBPD, EIF4E2, HS2ST1, AGPS, PGK1, ATIC, ETNK1, LG2, NCE2, MARCH8, CNOT4, RNF8, PSMA5, DPF2, AMMECR1, KIAA1287, LOC144233, LOC286505, PNAS-4, FLJ20530, HUMPD3, GC45564, CAP350, ETAA16, HAN11, DNAPTP6, C7orf25, FLJ37953, FLJ10587, C7orf36, ELP4, NDEL1, NPD014, KFZP564D172, FAM53C, IER5, LOC255783, KIAA0146, KIAA0792, LOC439994, LOC283481, CG018, LOC130576, NGFRAP1L1, KIAA1217, 4orf7, C21 orf86, C9orf64, FLJ13456, FLJ12806, FLJ39370, GATS, CCDC92, FMNL2, ARID1B, ZFHX1B, SSBP2, ARID5B, LOC157381, KPNA3, ARHGAP24, CCND2, VIM, MYCBP2, GAS7, TRAM2, BASP1, FOXO1A, POLR2A, PER1, DDIT4, CD97, BIN1, SH2B3, DDB2, EMP3, NDST2, CHST2, NT5E, PDE4A, CPS1, PTGS1, GGCX, IRF5, ZBTB16, MAP4K4, CHST7, KLF12, NFRKB, PSD, FKSG49, NIFUN, FYN, ZMYM2, CACNA1G, SLC25A16, FLII, EIF1, SEPT6, NPAC, PHF15, NUP188, ABR, CNR1, LOC283824, FSTL4, DNM1, APOBEC3G, ATP2B1, SMPD1, SLC11A1, FXYD5, C14orf139, SH3BGRL3, LUZP1, ZNF335, SH3 KBP1, MST150, PRO1073, LOC388397, FKBP5, HIPK2, DTX1, ST3GAL2, ADAMTS7, TNRC6B, CYGB, SDHAL1, LOC572558, TRIO, FRAS1, KIAA1632, POLS, EBF, MAML2, PTPRA, PLEKHG2, DYM, SOX6, ARHGEF2, ZCCHC6, PPP3CA, FAM70B, TMED5, FLJ43663, HPS1, MEF2A, ST3GAL5, SMYD3, KLF7, LOC200230, RERE, QKI, BICD1, CTNNB1, POU2F2, EIF4ENIF1, BTG1, TCHP, FLJ90709, PCTK2, KIAA1600, B7-H4, PDE4DIP, KIAA0194, HOM-TES; and
- (a) BGN, BIN1, CDH11, CDH2, CHN1, COL1A2, COL3A1, COL5A2, CREB3L1, CTGF, CYBRD1, DCN, DLC1, DNAJB4, DPT, ENPP2, FBLN1, FBLN5, FBN1, GREM1, HAS2, IGFBP3, IGFBP4, MLPH, MME, MMP2, MYL9, NEBL, NID1, NR2F1, OLFML3, PCOLCE, PMP22, POSTN, PPAP2B, PRR16, PRRX1, PTX3, PVRL3, RGL1, RGS4, ROR1, SEMA5A, TNFAIP6, TRAM2, WNT5A, ZEB1, ANXA6, CITED2, COL1A1, COL6A1, CYP1B1, DAB2, DOCK10, EML1, GNG11, IER3, ITGBL1, KCNMA1, LRIG1, LUM, MAP1B, NID2, PCDH9, PRKCA, PTGER4, PTGFR, SDC2, SERPINF1, SNED1, SYT11, TFPI, TGFBR3, VCAN, ANKRD25, C10ORF56, C5ORF13, CCDC92, CDKN2C, COPZ2, DDR2, ECM1, ENOX1, FADS2, FAP, FILIP1L, FSTL1, HS3ST2, KRT81, LMCD1, LTBP1, LTBP2, MMP1, NPAC, NRP1, PDGFC, PLEKHC1, PPM1D, PTGER2, SCCPDH, SEPT9, SPOCK1, SRGN, STARD13, STC1, SYNC1, TAGLN, TBX3, TGFB1I1, THY1, TNS3, TPM1, TSHZ1, TUBA1A, UGDH, VIM, XYLT1, ZBTB38, MBP, ABLIM1, ALDH1A3, ALOX15B, ANXA3, AP1M2, ARTN, CA2, CA9, CAMK2B, CCND2, CDH1, CDH3, CDS1, COL17A1, CORO1A, CTSL2, CXADR, CYP27B1, DSC2, DSG3, DST, EPB41L4B, FGFBP1, FGFR3, FST, GJB3, GRHL2, HBEGF, HOOK1, IL18, IL1B, IRF6, ITGB4, JAG2, KLK10, KLK5, KLK7, KLK8, KRT15, KRT16, KRT17, KRT5, KRT6B, LEPREL1, MYO5C, NDRG1, NMU, PI3, RAB25, RLN2, RNF128, S100A14, S100A7, S100A8, SERPINB1, SERPINB2, SLC2A9, SLPI, SNX10, SPINT2, SPRR1A, SPRR1B, ST14, SYK, TMEM40, TRIM29, TSPAN1, UCHL1, VSNL1, ZBED2, AQP3, CBARA1, CD24, CLCA2, CNTNAP2, CSTA, DSC3, EPCAM, ESRP2, FXYD3, GJB5, GPR87, IL1A, LAMA3, LAMB3, MAP7, MAPK13, MMP9, MPZL2, NRG1, SCNN1A, SERPINB13, SLC7A5, SPINT1, ST6GALNAC2, TGFA, TP63, TPD52L1, VGLL1, SLC6A8, ABCA12, ANK3, ANXA8, ANXA8L1, ARHGAP25, ARHGAP8, BDKRB2, BIK, C10ORF10, C10ORF116, C1ORF116, C20ORF19, C6ORF105, CELSR2, CKMT1A, CKMT1B, CLDN1, CST6, CYP4F11, E2F5, ELMO3, EPHA1, EVA1, EXPH5, F11R, FAT2, FGFR2, FLJ12684, FLJ20366, FZD3, GLS2, GNAL, IFI30, IGFBP2, IL1RN, IL4R, IRX4, JUP, KCNK1, KIAA0040, KIAA0888, KIAA1815, KRT14, KRT18, LAD1, LGALS7, LOC553158, LOC653562, LOC728113, LOC729884, LRRC1, LSR, MST1R, MYO1D, NAIP, OCLN, NEFM, NUP62CL, PERP, PLS1, POLR3G, PRKCH, PRRG4, PRSS8, PTPN3, RBM35A, RBM35B, RHBDF2, RHOD, RPS6KA1, RTEL1, SAA1, SAA1, SAA2, SH2D3A, SLC6A10P, SMPDL3B, SNCA, STAC, STAP2, TACSTD1, THBD, TMEM30B, TMPRSS11E, TMPRSS4, TNFRSF6B, TP73L, WWC1, XDH, ZNF165; and
- (b) BGN, BIN1, CDH11, CDH2, CHN1, COL1A2, COL3A1, COL5A2, CREB3L1, CTGF, CYBRD1, DCN, DLC1, DNAJB4, DPT, ENPP2, FBLN1, FBLN5, FBN1, GREM1, HAS2, IGFBP3, IGFBP4, MLPH, MME, MMP2, MYL9, NEBL, NID1, NR2F1, OLFML3, PCOLCE, PMP22, POSTN, PPAP2B, PRR16, PRRX1, PTX3, PVRL3, RGL1, RGS4, ROR1, SEMA5A, TNFAIP6, TRAM2, WNT5A, ZEB1, ANXA6, CITED2, COL1A1, COL6A1, CYP1B1, DAB2, DOCK10, EML1, GNG11, IER3, ITGBL1, KCNMA1, LRIG1, LUM, MAP1B, NID2, PCDH9, PRKCA, PTGER4, PTGFR, SDC2, SERPINF1, SNED1, SYT11, TFPI, TGFBR3, VCAN, ANKRD25, C10ORF56, C5ORF13, CCDC92, CDKN2C, COPZ2, DDR2, ECM1, ENOX1, FADS2, FAP, FILIP1L, FSTL1, HS3ST2, KRT81, LMCD1, LTBP1, LTBP2, MMP1, NPAC, NRP1, PDGFC, PLEKHC1, PPM1D, PTGER2, SCCPDH, SEPT9, SPOCK1, SRGN, STARD13, STC1, SYNC1, TAGLN, TBX3, TGFB1I1, THY1, TNS3, TPM1, TSHZ1, TUBA1A, UGDH, VIM, XYLT1, ZBTB38; and
- (c) ENPP2, CITED2, ZEB1, NID2, SEMA5A, DAB2, KCNMA1, PTX3, PCDH9, CREB3L1, BGN, ROR1, DLC1, GNG11, CDH11, NR2F1, NR2F2, PRR16, MYL9, DOCK10, LRIG1, IER3, EML1, NEBL, RGL1, MLPH, TRAM2, DNAJB4, FBLN5, RGS4, HAS2, ITGBL1, IGFBP4, BIN1, DPT, PCOLCE, GREM1, PPAP2B, CDH2, PMP22, LUM, CHN1, CYP1B1, MME, WNT5A, POSTN, MMP2, CTGF, FBLN1, IGFBP3, DCN, PRRX1, ANXA6, PVRL3, MAP1B, TNFAIP6, CYBRD1, FBN1, NID1, OLFML3, SNAI1, SNAI2, TAGLN, SYNC, TUBA1A, PPM1D, TWIST1, FN1, TGFBR3, SERPINF1, UGDH, SRGN, FAP, PTGER4, VIM, PRKCA, FSTL1, MMP1, NRP1, FILIP1L, SCCPDH, LTBP2, XYLT1, HS3ST2, SYT11, TSHZ1, THY1, SEPT9, S100A4, CCDC92, TNS3, ENOX1, TGFB1I1, ZEB2, LMCD1, PDGFC, ECM1, TFPI, TBX3, DDR2, PTGER2, CDKN2C, VCAN, CD44, STARD13, SNED1, ZBTB38, SDC2, TPM1, COPZ2, STC1, CDH1, KRT5, KRT6B, CD24, EPCAM, ESRP1, CLDN3, CLDN4, ERBB3, SPOCK1, FERMT2, GLYR1, LTBP1, FADS2, KANK2, PTGFR, ARTN, SLC7A5, EPB41L4B, KRT15, CDH3, TPD52L1, ANXA3, PI3, NDRG1, DSG3, S100A14, S100S7, KLK8, S100A8, PTX3, SPINT2, FXYD3, KRT16, RAB25, LGALS7, DSC3, COL5A2, ITGB4, TMEM40, SPRR1A, AP1M2, SERPINB13, TP73L; and
- (d) CYBRD1, ARTN, CITED2, SLC7A5, CHN1, CTGF, CSTA, MYL9, EPB41L4B, KRT15, LRIG1, CDH3, TPD52L1, ANXA3, OLFML3, CORO1A, PI3, NDRG1, DSG3, PMP22, S100A14, S100S7, KLK8, S100A8, PTX3, SPINT2, IER3, FXYD3, KRT16, TGFBR3, KRT5, RAB25, SDC2, LUM, LGALS7, KRT6B, CD24, COL1A2, GNG11, DSC3, COL5A2, RGS4, ITGB4, TMEM40, SPRR1A, AP1M2, SERPINB13, SNED1, LEPREL1, TRAM2, TP73L, and PAP2B; and
- (e) CYBRD1, ARTN, CITED2, SLC7A5, CHN1, CTGF, CSTA, MYL9, EPB41L4B, KRT15, LRIG1, CDH3, TPD52L1, ANXA3, OLFML3, CORO1A; and
- (f) PI3, NDRG1, DSG3, PMP22, S100A14, S100S7, KLK8, S100A8, PTX3, SPINT2, IER3, FXYD3, KRT16, TGFBR3; and
- (g) KRT5, RAB25, SDC2, LUM, LGALS7, KRT6B, CD24, COL1A2, GNG11, DSC3, COL5A2; and
- (h) RGS4, ITGB4, TMEM40, SPRR1A, AP1M2, AERPINB13, SNED1, LEPREL1, TRAM2, TP73L, and PAB2B; and
- (i) CYBRD1, ARTN, KRT15, ITGB4, RGS4, SPRR1B, SLC7A5, ANXA3, IL18, AP1M2, CHN1, OLFML3, ZBED2, LEPREL1, ABLIM1, CDH3, DNAJB4, TRAM2, UCHL1, CTGF; and
  - (i) CYBRD1, ARTN, KRT15, ITGB4, RGS4, SPRR1B, SLC7A5, ANXA3, IL18, AP1M2; and
  - (ii) CYBRD1, ARTN, KRT15, ITGB4, RGS4; and
- (j) CYBRD1, RGS4, CDH3, CHN1, SLC7A5, PRRX1, CITED2, COL5A2, ARTN, ZBED2, SNED1, ITGB4, LUM, PVRL3, OLFML3, BIN1, CCND2, DAB2, ANXA3, IL18; and
  - (i) CYBRD1, RGS4, CDH3, CHN1, SLC7A5, PRRX1, CITED2, COL5A2, ARTN, ZBED2; and
  - (ii) CYBRD1, RGS4, CDH3, CHN1, SLC7A5; and
- (k) CYBRD1, ARTN, RGS4, CITED2, SLC7A5, ITGB4, CHN1, PI3, TMEM40, NDRG1, SPRR1A, PCDH9, CTGF, MAP1B, DSG3, AP1M2, FBLN5, SERPINB13, PMP22, CSTA; and
  - (i) CYBRD1, ARTN, RGS4, CITED2, SLC7A5, ITGB4, CHN1, PI3, TMEM40, NDRG1; and
  - (ii) CYBRD1, ARTN, RGS4, CITED2, SLC7A5; and

(ii) said second set of genes comprises or consists of:
AXL, NOTCH3, GPR30, PROCR, PAX2, PI3KA, CD44, CD133, ALDH1, BMI1, KRT19, NRP1, MSI1, MTOR, JAG1, p38, ESA1, ZEB1, TWIST1, FOXC2, CDH2, ZEB2, FOXC1, SNAI1, SNAI2, TWIST2, GSC, TCF3, CD44, VIM, FN1, MLPH1, ARHGEF11; and
(iii) said third set of genes comprises or consists of:
At least one gene from said first set of genes and at least one of:
AXL, NOTCH3, GPR30, PROCR, PAX2, PI3KA, CD44, CD133, ALDH1, BMI1, KRT19, NRP1, MSI1, MTOR, JAG1, p38, ESA1, ZEB1, TWIST1, FOXC2, CDH2, ZEB2, FOXC1, SNAI1, SNAI2, TWIST2, GSC, TCF3, CD44, VIM, FN1, MLPH1, ARHGEF11; and
(iv) said fourth set of genes comprises or consists of:
At least two genes from said first set of genes and at least one of: AXL, NOTCH3, GPR30, PROCR, PAX2, PI3KA, CD44, CD133, ALDH1, BMI1, KRT19, NRP1, MSI1, MTOR, JAG1, p38, ESA1, ZEB1, TWIST1, FOXC2, CDH2, ZEB2, FOXC1, SNAI1, SNAI2, TWIST2, GSC, TCF3, CD44, VIM, FN1, MLPH1, and ARHGEF11; and
(v) said fifth set of genes comprises or consists of:
beta catenin, FZD1, FZD2, FZD3, FZD4, FZD5, FZD6, FZD7, FZD8, FZD9, FZD10, LRP5, LRP6, WNT1, WNT2, WNT2B, WNT3, WNT3A, WNT4 WNT5A, WNT5B, WNT6, WNT7A, WNT7B, WNT8A, WNT8B, WNT9A, WNT9B, WNT10A, WNT10B, WNT11, WNT16, SRFP1, SRFP2, FRZB, SFRP4, SFRP5, DKK1, DKK2, DKK3, DKK4, WIF1, AKT2, PI3KA, Disheveled, Smoothened, Axin1, Axin2; and
(vi) said sixth set of genes comprises or consists of:
At least one gene from said first set of genes and at least one of:
beta catenin, FZD1, FZD2, FZD3, FZD4, FZD5, FZD6, FZD7, FZD8, FZD9, FZD10, LRP5, LRP6, WNT1, WNT2, WNT2B, WNT3, WNT3A, WNT4 WNT5A, WNT5B, WNT6, WNT7A, WNT7B, WNT8A, WNT8B, WNT9A, WNT9B, WNT10A, WNT10B, WNT11, WNT16, SRFP1, SRFP2, FRZB, SFRP4, SFRP5, DKK1, DKK2, DKK3, DKK4, WIF1, AKT2, PI3KA, Disheveled, Smoothened, Axin1, Axin2; and
(vii) said seventh set of genes comprises or consists of:
TGFbeta1, TGFbeta2, TGFbeta3, ALK1, ALK2, ALK3, ALK4, ALK5, ALK6, ALK7, TGFbetaR-II, BMPR-II, ACTR-IIA, ACTR-IIB, AMHR-II, SMAD2, SMAD3; and
(viii) said eighth set of genes comprises or consists of:
At least one gene from said first set of genes and at least one of:
TGFbeta1, TGFbeta2, TGFbeta3, ALK1, ALK2, ALK3, ALK4, ALK5, ALK6, ALK7, TGFbetaR-II, BMPR-II, ACTR-IIA, ACTR-IIB, AMHR-II, SMAD2, SMAD3; and
(ix) said ninth set of genes comprises or consists of:
S100A4, SDC2, CYP1B1, FERMT2, CYBRD1, SNAI2, CD44, BIN1, PMP22, CTGF, IGFBP3, ANXA6, MAP1B, TAGLN, ENPP2, FBLN5, PCOLCE, FN1, VIM, PPAP2B, SEM5A, VCAN, NID2, NID1, HAS2, PRRX1, TNFAIP6, CLDN4, FADS2, DDIT4, GNG11, PER1, ISCU, KPNA3, SLC44A1, FKBP5, NR2F1, MLPH, DOCK10, RGS4, IGFBP4; and
(x) said tenth set of genes comprises or consists of:
S100A4, SDC2, CYP1B1, FERMT2, CYBRD1, SNAI2, CD44, BIN1, ENPP2, FBLN5, PCOLCE, FN1, VIM, PPAP2B, SEM5A, VCAN, CLDN4, FADS2, DDIT4, GNG11, PER1, ISCU, KPNA3, SLC44A1, FKBP5, IGFBP4; and
(xi) said eleventh set of genes comprises or consists of:
SDC2, CYBRD1, SNAI2, PMP22, CTGF, IGFBP3, ANXA6, MAP1B, TAGLN, PPAP2B, HAS2, NRF2, MLPH, DOCK10, RGS4; and
(xii) said twelfth set of genes comprises or consists of:
FERMT2, BIN1, PMP22, CHN1, CTGF, IGFBP3, NID2, CLDN4, CYBRD1, PRR16, ANXA6, PCOLCE, PPAP2B, HAS2, CDH11, MLPH, DOCK10, RGS4, PVRL3, NID2, SLC44A1, FKBP5; and
(xiii) said thirteenth set of genes comprises or consists of:
FERMT2, BIN1, PMP22, CHN1, CTGF, IGFBP3, NID2, CLDN4; and
(xiv) said fourteenth set of genes comprises or consists of:
HAS2, BIN1, PCOLCE, FERMT2, CTGF, IGFBP3, NID2, SLC44A1, FKBP5, and MLPH; and

- optionally, administering the agent to the subject.

In some embodiments, the determining the level of gene expression product comprises determining the level of RNA expression of each gene of said plurality of genes. In an embodiment, the level of gene expression is a function of the level of RNA expression of each gene of said plurality of genes. In an embodiment, the level of RNA expression is acquired. In an embodiment, the level of RNA expression of said plurality of genes is assayed. In an embodiment, the level of RNA expression is assayed by detecting an RNA product, e.g., mRNA of said sample. In an embodiment, the level of RNA expression is assayed by a hybridization based method, e.g., hybridization with a probe that is specific for said RNA product. In an embodiment, the level of RNA expression is assayed by; applying said sample, or the mRNA isolated from, or amplified from; said sample, to a nucleic acid microarray, or chip array. In an embodiment, the level of RNA expression is assayed by microarray. In an embodiment, the level of RNA expression is assayed by a polymerase chain reaction (PCR) based method, e.g., qRT-PCR. In an embodiment, the level of RNA expression is assayed by a sequencing based method. In an embodiment, the level of RNA expression is assayed by quantitative RNA sequencing. In an embodiment, the level of RNA expression is assayed by RNA in situ hybridization. In an embodiment, the level of RNA expression is assayed in the whole subject sample. In an embodiment, the level of RNA expression is assayed in a subregion of the subject sample, e.g., subregions of a tissue sample.
In some embodiments, the determining the level of gene expression product comprises determining the level of protein expression of each gene of said plurality of genes. In an embodiment, the level of protein expression is acquired. In an embodiment, the level of protein expression is assayed. In an embodiment, the level of protein expression is assayed by detecting a protein product. In an embodiment, the level of protein expression is assayed using antibodies selective for said protein product. In an embodiment, the level of protein expression is assayed by an immunohistochemistry technique. In an embodiment, the level of protein expression is assayed by an immunohistochemistry technique, using antibodies specific for said protein product. In an embodiment, the level of protein expression is assayed by an immunoassay, e.g., Western blot, enzyme linked immunosorbant assay (ELISA). In an embodiment, the level of protein expression is assayed by an immunoassay specific for said protein. In an embodiment, levels of gene expression are assessed using protein activity assays, such as functional assays. In an embodiment, the level of protein expression is assayed in the whole subject sample. In an embodiment, the level of protein expression is assayed in a subregion of the subject sample, e.g., subregions of a tissue sample.
In some embodiments, the method further comprises determining the level of gene expression product in a cell. In some embodiments, the determining the level of gene expression product in a cell comprises: contacting the cell with an agent; determining the level of gene expression product; and comparing the level of gene expression product to an appropriate control.
In some embodiments, the subject sample is a sample described herein, e.g., blood, urine, or tissue sample. In an embodiment, the subject sample is a tissue sample, e.g., biopsy. In an embodiment, the subject sample is a bodily fluid, e.g., blood, plasma, urine, saliva, sweat, tears, semen, or cerebrospinal fluid. In an embodiment, the subject sample is a bodily product, e.g., exhaled breath. In an embodiment, said subject sample is a tissue sample, wherein said tissue sample is derived from fixed tissue, paraffin embedded tissue, fresh tissue, or frozen tissue. In an embodiment, said subject sample is a tissue sample, wherein said tissue sample is fixed tissue, paraffin embedded tissue, fresh tissue, or frozen tissue.
In some embodiments the subject has cancer, e.g., a cancer described herein, e.g., breast cancer. The cancer can include cancers characterized as comprising cancer stem cells, cancer associated mesenchymal cells, or tumor initiating cancer cells. The cancer can include cancers that have been characterized as being enriched with cancer stem cells, cancer associated mesenchymal cells, or tumor initiating cancer cells. Exemplary cancers include epithelial cancers, breast, lung, pancreatic, colorectal, prostate, head and neck, melanoma, acute myelogenous leukemia, and glioblastoma. Exemplary breast cancers include triple negative breast cancer, basal-like breast cancer, claudin-low breast cancer, invasive, inflammatory, metaplastic, and advanced Her-2 positive or ER-positive cancers resistant to therapy. Other cancers include but are not limited to, brain, abdominal, esophagus, gastrointestinal, glioma, liver, tongue, neuroblastoma, osteosarcoma, ovarian, retinoblastoma, Wilm's tumor, multiple myeloma, skin, lymphoma, blood, retinal, acute lymphoblastic leukemia, bladder, cervical, kidney, endometrial, meningioma, lymphoma, skin, uterine, lung, non small cell lung, nasopharyngeal carcinoma, neuroblastoma, solid tumor, hematologic malignancy, leukemia, squamous cell carcinoma, testicular, thyroid, mesothelioma, brain vulval, sarcoma, intestine, oral, T cell leukemia, endocrine, salivary, spermatocytic seminoma, sporadic medulalry thyroid carcinoma, non-proliferating testes cells, cancers related to malignant mast cells, non-Hodgkin's lymphoma, and diffuse large B cell lymphoma.
The cancer can be a primary tumor, i.e., located at the anatomical site of tumor growth initiation. The cancer can also be metastatic, i.e., appearing at least a second anatomical site other than the anatomical site of tumor growth initiation. The cancer can be a recurrent cancer, i.e., cancer that returns following treatment, and after a period of time in which the cancer was undetectable. The recurrent cancer can be anatomically located locally to the original tumor, e.g., anatomically near the original tumor; regionally to the original tumor, e.g., in a lymph node located near the original tumor; or distantly to the original tumor, e.g., anatomically in a region remote from the original tumor.
Also described herein are in vitro methods and assays. In one aspect described herein are in vitro methods and assays of determining if a subject is a potential candidate for treatment with an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells, the method comprising determining the level of gene expression product of a plurality of genes selected from a first and/or second and/or third and/or fourth and/or fifth and/or sixth set of genes, in a subject sample, wherein:

- (i) said first set of genes comprises or consists of:
- S100A4, SDC2, CYP1B1, FERMT2, CYBRD1, SNAI2, CD44, BIN1, PMP22, CTGF, IGFBP3, ANXA6, MAP1B, TAGLN, ENPP2, FBLN5, PCOLCE, FN1, VIM, PPAP2B, SEM5A, VCAN, NID2, NID1, HAS2, PRRX1, TNFAIP6, CLDN4, FADS2, DDIT4, GNG11, PER1, ISCU, KPNA3, SLC44A1, FKBP5, NR2F1, MLPH, DOCK10, RGS4, IGFBP4; and
- (ii) said second set of genes comprises or consists of:
- S100A4, SDC2, CYP1B1, FERMT2, CYBRD1, SNAI2, CD44, BIN1, ENPP2, FBLN5, PCOLCE, FN1, VIM, PPAP2B, SEM5A, VCAN, CLDN4, FADS2, DDIT4, GNG11, PER1, ISCU, KPNA3, SLC44A1, FKBP5, IGFBP4; and
- (iii) said third set of genes comprises or consists of:
- SDC2, CYBRD1, SNAI2, PMP22, CTGF, IGFBP3, ANXA6, MAP1B, TAGLN, PPAP2B, HAS2, NRF2, MLPH, DOCK10, RGS4; and
- (iv) said fourth set of genes comprises or consists of:
- FERMT2, BIN1, PMP22, CHN1, CTGF, IGFBP3, NID2, CLDN4, CYBRD1, PRR16, ANXA6, PCOLCE, PPAP2B, HAS2, CDH11, MLPH, DOCK10, RGS4, PVRL3, NID2, SLC44A1, FKBP5; and
- (v) said fifth set of genes comprises or consists of:
- FERMT2, BIN1, PMP22, CHN1, CTGF, IGFBP3, NID2, CLDN4; and
- (vi) said sixth set of genes comprises or consists of:
- HAS2, BIN1, PCOLCE, FERMT2, CTGF, IGFBP3, NID2, SLC44A1, FKBP5, and MLPH; and
- optionally, administering the agent to the subject.

In some embodiments, the determining the level of gene expression product comprises determining the level of RNA expression of each gene of said plurality of genes. In an embodiment, the level of gene expression is a function of the level of RNA expression of each gene of said plurality of genes. In an embodiment, the level of RNA expression is acquired. In an embodiment, the level of RNA expression of said plurality of genes is assayed. In an embodiment, the level of RNA expression is assayed by detecting an RNA product, e.g., mRNA of said sample. In an embodiment, the level of RNA expression is assayed by a hybridization based method, e.g., hybridization with a probe that is specific for said RNA product. In an embodiment, the level of RNA expression is assayed by; applying said sample, or the mRNA isolated from, or amplified from; said sample, to a nucleic acid microarray, or chip array. In an embodiment, the level of RNA expression is assayed by microarray. In an embodiment, the level of RNA expression is assayed by a polymerase chain reaction (PCR) based method, e.g., qRT-PCR. In an embodiment, the level of RNA expression is assayed by a sequencing based method. In an embodiment, the level of RNA expression is assayed by quantitative RNA sequencing. In an embodiment, the level of RNA expression is assayed by RNA in situ hybridization. In an embodiment, the level of RNA expression is assayed in the whole subject sample. In an embodiment, the level of RNA expression is assayed in a subregion of the subject sample, e.g., subregions of a tissue sample.
In some embodiments, the determining the level of gene expression product comprises determining the level of protein expression of each gene of said plurality of genes. In an embodiment, the level of protein expression is acquired. In an embodiment, the level of protein expression is assayed. In an embodiment, the level of protein expression is assayed by detecting a protein product. In an embodiment, the level of protein expression is assayed using antibodies selective for said protein product. In an embodiment, the level of protein expression is assayed by an immunohistochemistry technique. In an embodiment, the level of protein expression is assayed by an immunohistochemistry technique, using antibodies specific for said protein product. In an embodiment, the level of protein expression is assayed by an immunoassay, e.g., Western blot, enzyme linked immunosorbant assay (ELISA). In an embodiment, the level of protein expression is assayed by an immunoassay specific for said protein. In an embodiment, levels of gene expression are assessed using protein activity assays, such as functional assays. In an embodiment, the level of protein expression is assayed in the whole subject sample. In an embodiment, the level of protein expression is assayed in a subregion of the subject sample, e.g., subregions of a tissue sample.
In some embodiments, the method further comprises determining the level of gene expression product in a cell. In some embodiments, the determining the level of gene expression product in a cell comprises: contacting the cell with an agent; determining the level of gene expression product; and comparing the level of gene expression product to an appropriate control.
In some embodiments, the subject sample is a sample described herein, e.g., blood, urine, or tissue sample. In an embodiment, the subject sample is a tissue sample, e.g., biopsy. In an embodiment, the subject sample is a bodily fluid, e.g., blood, plasma, urine, saliva, sweat, tears, semen, or cerebrospinal fluid. In an embodiment, the subject sample is a bodily product, e.g., exhaled breath. In an embodiment, said subject sample is a tissue sample, wherein said tissue sample is derived from fixed tissue, paraffin embedded tissue, fresh tissue, or frozen tissue. In an embodiment, said subject sample is a tissue sample, wherein said tissue sample is fixed tissue, paraffin embedded tissue, fresh tissue, or frozen tissue.
In some embodiments the subject has cancer, e.g., a cancer described herein, e.g., breast cancer. The cancer can include cancers characterized as comprising cancer stem cells, cancer associated mesenchymal cells, or tumor initiating cancer cells. The cancer can include cancers that have been characterized as being enriched with cancer stem cells, cancer associated mesenchymal cells, or tumor initiating cancer cells. Exemplary cancers include epithelial cancers, breast, lung, pancreatic, colorectal, prostate, head and neck, melanoma, acute myelogenous leukemia, and glioblastoma. Exemplary breast cancers include triple negative breast cancer, basal-like breast cancer, claudin-low breast cancer, invasive, inflammatory, metaplastic, and advanced Her-2 positive or ER-positive cancers resistant to therapy. Other cancers include but are not limited to, brain, abdominal, esophagus, gastrointestinal, glioma, liver, tongue, neuroblastoma, osteosarcoma, ovarian, retinoblastoma, Wilm's tumor, multiple myeloma, skin, lymphoma, blood, retinal, acute lymphoblastic leukemia, bladder, cervical, kidney, endometrial, meningioma, lymphoma, skin, uterine, lung, non small cell lung, nasopharyngeal carcinoma, neuroblastoma, solid tumor, hematologic malignancy, leukemia, squamous cell carcinoma, testicular, thyroid, mesothelioma, brain vulval, sarcoma, intestine, oral, T cell leukemia, endocrine, salivary, spermatocytic seminoma, sporadic medulalry thyroid carcinoma, non-proliferating testes cells, cancers related to malignant mast cells, non-Hodgkin's lymphoma, and diffuse large B cell lymphoma.
The cancer can be a primary tumor, i.e., located at the anatomical site of tumor growth initiation. The cancer can also be metastatic, i.e., appearing at least a second anatomical site other than the anatomical site of tumor growth initiation. The cancer can be a recurrent cancer, i.e., cancer that returns following treatment, and after a period of time in which the cancer was undetectable. The recurrent cancer can be anatomically located locally to the original tumor, e.g., anatomically near the original tumor; regionally to the original tumor, e.g., in a lymph node located near the original tumor; or distantly to the original tumor, e.g., anatomically in a region remote from the original tumor.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 depicts genes highly ranked from multiple statistical gene ranking methods. Genes listed are concordant between two of three statistical gene ranking methods: LASSO, STEPWISE, and CART. A one (1) indicates a gene that is one of the top 60 genes of the ranking algorithm. A zero (0) indicates a gene that is not one of the top 60 genes of the ranking algorithm, but is a top 60 gene of another ranking algorithm.

FIG. 2 depicts the concordance of the top 60 genes from three gene ranking methods. Three gene ranking methods were utilized to fit a gene rank order from EMT (epithelial-to-mesenchymal transition) and CSC genes. The three methods used were LASSO, STEPWISE, and CART. FIG. 2 illustrates the level of concordance between the three methods.

FIG. 3 illustrates the LASSO-20 gene model. The top twenty highly ranked genes from the LASSO gene ranking algorithm were used to distinguish ‘likely to recur’ and ‘no recurrence’ triple negative breast cancer patient subgroups. The left panel is a time to recurrence Kaplan-Meier plot for the Event-Free fraction from 0-7 years. As labeled, ‘Recurrence’ refers to ‘Likely to Recur’ and ‘No Recurrence’ refers to ‘Not Likely to Recur’. The right panel is a receiver operator characteristic (ROC) plot and calculation of Area-Under-Curve (AUC), as a projection of the relationship between the true positive rate and the false positive rate.

FIG. 4 illustrates the STEPWISE 20-gene model. The top twenty highly ranked genes from the STEPWISE gene ranking algorithm were used to distinguish ‘likely to recur’ and ‘no recurrence’ triple negative breast cancer patient subgroups. The left panel is a time to recurrence Kaplan-Meier plot for the Event-Free fraction from 0-7 years. As labeled, ‘Recurrence’ refers to ‘Likely to Recur’ and ‘No Recurrence’ refers to ‘Not Likely to Recur’. The right panel is a receiver operator characteristic (ROC) plot and calculation of Area-Under-Curve (AUC), as a projection of the relationship between the true positive rate and the false positive rate.

FIG. 5 illustrates the CART 20-gene model. The top twenty highly ranked genes from the CART gene ranking algorithm were used to distinguish ‘likely to recur’ and ‘no recurrence’ triple negative breast cancer patient subgroups. The left panel is a time to recurrence Kaplan-Meier plot for the Event-Free fraction from 0-7 years. As labeled, ‘Recurrence’ refers to ‘Likely to Recur’ and ‘No Recurrence’ refers to ‘Not Likely to Recur’. The right panel is a receiver operator characteristic (ROC) plot and calculation of Area-Under-Curve (AUC), as a projection of the relationship between the true positive rate and the false positive rate.

FIG. 6 illustrates the addition of driver genes to base gene models [Add-One-In]. The table lists candidate genes that are tested in a base gene model with 4 other genes (5-gene models). Genes are sorted by p-value improvement, when tested for their prediction of ‘recurrence’ versus ‘no recurrence’ sub grouping of triple negative breast cancer patients, where biopsies were isolated prior to treatment. High performing ‘add-in’ genes (p-value<1e−5) are shown as ̂̂genes, ** genes that are ‘positive control’ (Ranked top 5 genes via LASSO).

FIG. 7 illustrates recurrence plots for driver genes designated as Group A. Five gene models were tested for their prediction of ‘recurrence’ versus ‘no recurrence’ sub grouping of triple negative breast cancer patients, where biopsies were isolated prior to treatment. The figure depicts ‘time to recurrence’ Kaplan-Meier display for the ‘event-free fraction’ from 0-7 years. As labeled, ‘Recurrence’ refers to ‘Likely to Recur’ and ‘No Recurrence’ refers to ‘Not Likely to Recur’. The 5-gene models were constructed with a base model of 4 genes and addition of a single candidate driver gene. Driver genes were: MSI1, NRP1, TWIST1, BMI1, AXL, and ZEB1. Solid Lines depict the addition of a driver gene. Dotted Lines depict the addition of a positive control gene.

FIG. 8 illustrates recurrence plots for driver genes designated as Group B. Five gene models were tested for their prediction of ‘recurrence’ versus ‘no recurrence’ sub grouping of triple negative breast cancer patients, where biopsies were isolated prior to treatment. The figure depicts ‘time to recurrence’ Kaplan-Meier display for the ‘event-free fraction’ from 0-7 years. The 5-gene models were constructed with a base model of 4 genes and addition of a single candidate driver gene. Driver genes were: CD44, KRT19, PROCR, FOXC2, and NOTCH3. Solid Lines depict the addition of a driver gene. Dotted Lines depict the addition of a positive control gene.

FIG. 9 illustrates a recurrence plot for driver genes designated as a CSC group. A CSC gene model was tested for its prediction of ‘recurrence’ versus ‘no recurrence’ sub grouping of triple negative breast cancer patients, where biopsies were isolated prior to treatment. FIG. 9 depicts ‘time to recurrence’ Kaplan-Meier display for the ‘event-free fraction’ from 0-7 years. The nine CSC candidate genes evaluated were NOTCH3, PROCR, BMI1, KRT19, MSI1, NRP1, JAG1, ALDH1A1, and HTATIP. The p-value between the ‘recurrence’ and ‘no recurrence’ subgroups of patients was p=5.77e−4.

FIG. 10 illustrates a recurrence plot for a 5-gene CSC model. A five gene model was tested for its prediction of ‘recurrence’ versus ‘no recurrence’ subgroups of triple negative breast cancer patients, where biopsies were isolated prior to treatment. FIG. 10 depicts ‘time to recurrence’ Kaplan-Meier display for the ‘event-free fraction’ from 0-7 years. The genes evaluated in the model were NRP1, BMI1, JAG1, MSI1, and HTATIP. The p-value between the ‘recurrence’ and ‘no recurrence’ subgroups of patients was p=1.04e−3.

FIG. 11 illustrates the relative expression of candidate cancer stem cell and tumor initiating cancer cell genes, comparing tumorspheres with two-dimensional cell culture. The fold change in gene expression was measured by Q-PCR for RNA samples isolated from Tumorspheres or from two-dimensional cell culture of human breast cancer cell lines, MCF7, SUM159, and MDA-MB-231. The genes KRT5, SERPINF1, S100A4, RGL1, FAP, TGFBR3, FBLN5, MLPH, FADS2, TFPI, PPAP2B, SEPT9, CYBRD1, UGDH, TRAM2, PCDH9, and DLC1 were evaluated in this analysis. Plots show the fold change of Tumorsphere/2D culture on a log 2 scale for the three cell lines.

FIG. 12 illustrates a Time to Recurrence plot for 9 genes validated from tumorsphere culture. A Kaplan-Meier display for the ‘event-free fraction’ from 0-7 years is shown. As labeled, ‘Recurrence’ refers to ‘Likely to Recur’ and ‘No Recurrence’ refers to ‘Not Likely to Recur’. The genes in the model were CYBRD1, FAP, PPAP2B, TRAM2, TFPI, PCDH9, RGL1, FADS2, SERPINF1, DLC1, SEP9, MLPH, S100A4, FBLN5, and TGFBR3. A combined Gene Value was assigned that separates the patients into two groups. The p-value between the ‘Recurrence’ and ‘No recurrence’ subgroups of patients was p=6.83e−08.

FIG. 13 illustrates a recurrence plot for 5 genes validated from tumorsphere culture. The five gene model was tested for its prediction of ‘recurrence’ versus ‘no recurrence’ sub grouping of triple negative breast cancer patients, where biopsies were isolated prior to treatment. The figure depicts ‘time to recurrence’ Kaplan-Meier display for the ‘event-free fraction’ from 0-7 years. As labeled, ‘Recurrence’ refers to ‘Likely to Recur’ and ‘No Recurrence’ refers to ‘Not Likely to Recur’. The genes in the model were CYBRD1, FAP, PPAP2B, TRAM2, and TFPI. The p-value between the ‘Recurrence’ and ‘No recurrence’ subgroups of patients was p=2.19e−08.

FIG. 14 illustrates the upregulation of 11 cancer stem cell associated genes in sequential passaging of tumorspheres of SUM159 cells. The figure further illustrates the enhancement of the upregulation of the 11 cancer stem cell associated genes in paclitaxel treated SUM159 cell tumorspheres. The plot shows the percentage change in gene expression normalized to the expression level in the primary SUM159 tumorspheres.

FIG. 15 illustrates genes expressed in mesenchymal (M) tumor cells (i.e. basal breast cancer cell line), but are only expressed to a low or not expressed in epithelial (E) tumor cells (i.e. luminal breast cancer cell line) and fibroblast (F) non-tumor cells (i.e. fibroblasts). The values shown are the expression levels per gene per cell line, and the mean Expression level is indicated. These genes are designated as M-high, E-low, and Fibroblast-low. The example in the Box is one gene, EML1, where the Expression Level is shown for Basal, Luminal, and Fibroblast cell lines.

FIG. 16A illustrates a Recurrence plot for 19 genes designated as a M-high, E-low, and Fibroblast-low group. The 19 gene model was tested for its prediction of ‘recurrence’ versus ‘no recurrence’ sub grouping of triple negative breast cancer patients, where biopsies were isolated prior to treatment. The figure depicts ‘time to recurrence’ Kaplan-Meier display for the ‘event-free fraction’ from 0-7 years. As labeled, ‘Recurrence’ refers to ‘Likely to Recur’ and ‘No Recurrence’ refers to ‘Not Likely to Recur’. The genes in the model were CDH2, CHN1, CTGF, CYP1B1, DNAJB4, EML1, ENPP2, HAS2, MAP1B, NID1, PDGFC, PRR16, PRRX1, PVRL3, ROR1, SDC2, SNAI1, SNAI2, TNFAIP6, and VCAN. The p-value between the ‘Recurrence’ and ‘No recurrence’ subgroups of patients was p=1.9e−03. FIG. 16B is a receiver operator characteristic (ROC) plot and calculation of Area-Under-Curve (AUC), as a projection of the relationship between the true positive rate and the false positive rate, for the M-high, E-low, and Fibroblast-low 19 gene model. FIG. 16C illustrates a Recurrence plot for 6 genes designated as a M-high, E-low, and Fibroblast-low group. The 6 gene model was tested for its prediction of ‘recurrence’ versus ‘no recurrence’ sub grouping of triple negative breast cancer patients, where biopsies were isolated prior to treatment. The figure depicts ‘time to recurrence’ Kaplan-Meier display for the ‘event-free fraction’ from 0-7 years. As labeled, ‘Recurrence’ refers to ‘Likely to Recur’ and ‘No Recurrence’ refers to ‘Not Likely to Recur’. The genes in the 6 gene model were CHN1, CTGF, DNAJB4, PRRX1, PVRL3, and VCAN. The p-value between the ‘Recurrence’ and ‘No recurrence’ subgroups of patients was p=1.7e−04. FIG. 16D is a receiver operator characteristic (ROC) plot and calculation of Area-Under-Curve (AUC), as a projection of the relationship between the true positive rate and the false positive rate, for the M-high, E-low, and Fibroblast-low 6 gene model.

FIG. 17 illustrates the p-values associated with expression of significant genes (significance based on variance) over a large group of human breast cancer specimens. Twenty-five genes highly ranked by the variance criteria are shown to have a higher degree of variability as illustrated by p-value significance (below the indicated horizontal line, p<1.01).

FIG. 18 illustrates the polarity associated with expression of significant genes (significance based on polarity) over a large group of human breast cancer specimens. Genes with the highest and lowest Model Polarity are the ones that have the highest fraction of specimens that either upregulate or downregulate expression of the specified gene.

FIG. 19 illustrates a Time to Recurrence plot for an 18 gene model for candidate genes identified by Variance and Polarity. The 18 gene model was tested for its prediction of ‘recurrence’ versus ‘no recurrence’ subgroups of triple negative breast cancer patients, where biopsies were isolated prior to treatment. The left panel depicts ‘time to recurrence’ Kaplan-Meier display for the ‘event-free fraction’ from 0-7 years. The p-value between the ‘recurrence’ and ‘no recurrence’ subgroups of patients was p<0e+00. The right panel is a receiver operator characteristic (ROC) plot and calculation of Area-Under-Curve (AUC), as a projection of the relationship between the true positive rate and the false positive rate.

FIG. 20 illustrates a Time to Recurrence plot for a 12 gene model for candidate genes identified by Variance and Polarity. The 12 gene model was tested for its prediction of ‘recurrence’ versus ‘no recurrence’ subgroups of triple negative breast cancer patients, where biopsies were isolated prior to treatment. The left panel depicts ‘time to recurrence’ Kaplan-Meier display for the ‘event-free fraction’ from 0-7 years. The p-value between the ‘recurrence’ and ‘no recurrence’ subgroups of patients was p<0e+00. The right panel is a receiver operator characteristic (ROC) plot and calculation of Area-Under-Curve (AUC), as a projection of the relationship between the true positive rate and the false positive rate.

FIG. 21 illustrates a Time to Recurrence plot for a 6 gene model for candidate genes identified by Variance and Polarity. The 6 gene model was tested for its prediction of ‘recurrence’ versus ‘no recurrence’ subgroups of triple negative breast cancer patients, where biopsies were isolated prior to treatment. The left panel depicts ‘time to recurrence’ Kaplan-Meier display for the ‘event-free fraction’ from 0-7 years. The p-value between the ‘recurrence’ and ‘no recurrence’ subgroups of patients was p<1.22e−10. The right panel is a receiver operator characteristic (ROC) plot and calculation of Area-Under-Curve (AUC), as a projection of the relationship between the true positive rate and the false positive rate.

FIG. 22A illustrates a Time to Recurrence plot for a Tumorsphere gene model. The 6 gene model (CYBRD1, SERPINF1, FAP, PPAP2B, RGS4 and PRRX1) was tested for its prediction of ‘recurrence’ versus ‘no recurrence’ subgroups of triple negative breast cancer patients, where biopsies were isolated prior to treatment. The left panel depicts ‘time to recurrence’ Kaplan-Meier display for the ‘event-free fraction’ from 0-7 years. The p-value between the ‘recurrence’ and ‘no recurrence’ subgroups of patients was p=1.21e−07. FIG. 22B illustrates a receiver operator characteristic (ROC) plot and calculation of Area-Under-Curve (AUC), as a projection of the relationship between the true positive rate and the false positive rate, for the 6 gene model (CYBRD1, SERPINF1, FAP, PPAP2B, RGS4, and PRRX1). An AUC of 0.756 was observed. FIG. 22C illustrates a Time to Recurrence plot for a Tumorsphere 8-gene model combining CYBRD1, SERPINF1, FAP, PPAP2B with RGS4, PRRX1, FBXO21, and ID3. The 8 gene model was tested for its prediction of ‘recurrence’ versus ‘no recurrence’ subgroups of triple negative breast cancer patients where biopsies were isolated prior to chemotherapy treatment. The left panel depicts ‘time to recurrence’ Kaplan-Meier display for the ‘event-free fraction’ from 0-7 years. The p-value between the ‘recurrence’ and ‘no recurrence’ subgroups of patients was p=2.66e−12. FIG. 22B illustrates a receiver operator characteristic (ROC) plot and calculation of Area-Under-Curve (AUC), as a projection of the relationship between the true positive rate and the false positive rate, for the 8-gene model combining CYBRD1, SERPINF1, FAP, PPAP2B with RGS4, PRRX1, FBXO21, and ID3. An AUC was computed to 0.821 in Receiver Operator Curve analysis.

DETAILED DESCRIPTION

Certain terms are first defined. Additional terms are defined throughout the specification.
“Acquire” or “acquiring” as the terms are used herein, refer to obtaining possession of a physical entity, or a value, e.g., a numerical value, by “directly acquiring” or “indirectly acquiring” the physical entity or value. “Directly acquiring” means performing a process (e.g., performing a synthetic or analytical method) to obtain the physical entity or value. “Indirectly acquiring” refers to receiving the physical entity or value from another party or source (e.g., a third party laboratory that directly acquired the physical entity or value). Directly acquiring a physical entity includes performing a process that includes a physical change in a physical substance, e.g., a starting material. Exemplary changes include making a physical entity from two or more starting materials, shearing or fragmenting a substance, separating or purifying a substance, combining two or more separate entities into a mixture, performing a chemical reaction that includes breaking or forming a covalent or non-covalent bond. Directly acquiring a value includes performing a process that includes a physical change in a sample or another substance, e.g., performing an analytical process which includes a physical change in a substance, e.g., a sample, analyte, or reagent (sometimes referred to herein as “physical analysis”), performing an analytical method, e.g., a method which includes one or more of the following: separating or purifying a substance, e.g., an analyte, or a fragment or other derivative thereof, from another substance; combining an analyte, or fragment or other derivative thereof, with another substance, e.g., a buffer, solvent, or reactant; or changing the structure of an analyte, or a fragment or other derivative thereof, e.g., by breaking or forming a covalent or non-covalent bond, between a first and a second atom of the analyte; or by changing the structure of a reagent, or a fragment or other derivative thereof, e.g., by breaking or forming a covalent or non-covalent bond, between a first and a second atom of the reagent.
“Acquiring a sample” as the term is used herein, refers to obtaining possession of a sample, e.g., a tissue sample or nucleic acid sample, by “directly acquiring” or “indirectly acquiring” the sample. “Directly acquiring a sample” means performing a process (e.g., performing a physical method such as a surgery or extraction) to obtain the sample. “Indirectly acquiring a sample” refers to receiving the sample from another party or source (e.g., a third party laboratory that directly acquired the sample). Directly acquiring a sample includes performing a process that includes a physical change in a physical substance, e.g., a starting material, such as a tissue, e.g., a tissue in a human patient or a tissue that has was previously isolated from a patient. Exemplary changes include making a physical entity from a starting material, dissecting or scraping a tissue; separating or purifying a substance (e.g., a sample tissue or a nucleic acid sample); combining two or more separate entities into a mixture; performing a chemical reaction that includes breaking or forming a covalent or non-covalent bond. Directly acquiring a sample includes performing a process that includes a physical change in a sample or another substance, e.g., as described above. As used herein, a subject who is a “candidate” is a one likely to respond to a particular therapeutic regimen, relative to a reference subject or group of subjects. A “non-candidate” subject is one not likely to respond to a particular therapeutic regimen, relative to a reference subject or group of subjects.
The term “cancer stem cell” refers to a cell or group of cells in a tumor having stem-like progenitor properties.
The term “tumor initiating cancer cell” refers to a cell with stem-like properties and the ability to initiate a tumor upon introduction into a tissue.
The term “cancer associated mesenchymal cell” refers to a cell or cells in a tumor that have acquired or retained mesenchymal properties.
The term “anti-cancer stem cell agent” refers to an inhibitor or killer of cancer stem cells causing a reduction or elimination of these cells or a reduction in the ability of these cells to proliferative or to survive the treatment.
The term “agent that inhibits or kills cancer associated mesenchymal cells” refers to an inhibitor or killer of cancer mesenchymal cells causing a reduction or elimination of these cells or a reduction in the ability of these cells to proliferative or to survive the treatment.
The term “agent that inhibits or kills tumor initiating cancer cells” refers to an inhibitor or killer of cells with stem-like properties and the ability to initiate a tumor upon introduction into a tissue.
The term “agent that kills or inhibits cancer stem cells” refers to an inhibitor or killer of cells or a group of cells in a tumor having stem-like progenitor properties.
The term “anti-cancer agent” refers to an inhibitor of cancer initiation, growth, progression, or metastasis
The terms “cancer” and “tumor” are used interchangeably herein. These terms refer to the presence of cells possessing characteristics typical of cancer-causing cells, such as uncontrolled proliferation, immortality, metastatic potential, rapid growth and proliferation rate, and certain characteristic morphological features. Cancer cells are often in the form of a tumor, but such cells can exist alone within an animal, or can be a non-tumorigenic cancer cell, such as a leukemia cell. These terms include a solid tumor, a soft tissue tumor, or a metastatic lesion.
“Chemotherapeutic agent” means a chemical substance, such as a cytotoxic or cytostatic agent, that is used to treat a condition, particularly cancer. As used herein, “chemotherapy” and “chemotherapeutic” and “chemotherapeutic agent” are synonymous terms.
“Likely to” or “increased likelihood,” as used herein, refers to an increased probability that an item, object, thing or person will occur. Thus, in one example, a subject that is likely to respond to treatment with an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells; alone or in combination, has an increased probability of responding to treatment with the agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cell; alone or in combination, relative to a reference subject or group of subjects.
The term “location”, as used herein, refers to a zone of a sample defined by preselected criteria, such as morphology, histopathology, and other attributes. A zone of a tumor can be defined by a unique gene expression pattern of a set of preselected genes. A zone may be classified as containing a specific cell type or multiple cell types, e.g., a zone may be classified as a nodule of cancer stem cells; a nodule of cancer associated mesenchymal cells; a nodule of tumor initiating cancer cells; a zone of transition, e.g., an area between epithelial and mesenchymal features of a tumor region; or it may be a niche indicated by the presence of a particular cell type or class, e.g., mesenchymal cells, stromal cells, inflammatory cells, endothelial cells, etc.
“Unlikely to” or “decreased likelihood” refers to a decreased probability that an event, item, object, thing or person will occur with respect to a reference. Thus, a subject that is unlikely to respond to treatment with an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells; alone or in combination, has a decreased probability of responding to treatment with the agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells; alone or in combination, relative to a reference subject or group of subjects.
“Sequencing” a nucleic acid molecule requires determining the identity of at least one nucleotide in the molecule. The identity of less than all of the nucleotides in a molecule can be determined. The identity of a majority or all of the nucleotides in the molecule can be determined.
The terms “sample” and “subject sample” are used interchangeably herein. These terms refer to biological material obtained from a subject. The source of the sample can be solid tissue as from a fresh, frozen and/or preserved organ, tissue sample, biopsy, or aspirate; blood or any blood constituents; bodily fluids such as cerebral spinal fluid, amniotic fluid, peritoneal fluid or interstitial fluid; or cells from any time in gestation or development of the subject. The tissue sample can contain compounds that are not naturally intermixed with the tissue in nature such as preservatives, anticoagulants, buffers, fixatives, nutrients, antibiotics or the like. The sample can be preserved as a frozen sample or as formaldehyde- or paraformaldehyde-fixed paraffin-embedded (FFPE) tissue preparation. For example, the sample can be embedded in a matrix, e.g., an FFPE block or a frozen sample. The sample can also be a cell line, a cell line previously established, a cell line derived previously from a subject, etc.
The terms “treat” and “treatment” and “treatment regimen” and “therapeutic regimen” are used interchangeably herein. As used herein, the terms “treat” and “treatment” and “treatment regimen” and “therapeutic regimen” are defined as the application or administration of a compound, alone or in combination with, a second compound to a sample, e.g., a sample, or application or administration of the compound to an isolated tissue or cell, e.g., cell line, from a subject, e.g., a subject, who has a disorder (e.g., a disorder as described herein), a symptom of a disorder, or a predisposition toward a disorder, with the purpose to cure, heal, alleviate, relieve, alter, remedy, ameliorate, improve or affect the disorder, one or more symptoms of the disorder or the predisposition toward the disorder (e.g., to minimize at least one symptom of the disorder or to delay onset of at least one symptom of the disorder).
A “weighting factor” as used herein, refers to an element used as an adjustment factor for a specific value or group of similar values.
A subject that will “respond positively” or “respond favorably” as used herein, refers to a subject that will experience some degree of alleviation in one or more characteristics of a disease or disorder after receiving treatment with a therapeutic agent; and/or some degree of alleviation in one or more symptoms caused by a disease or disorder, after receiving treatment with a therapeutic agent.
A “responder” as used herein, is a subject that will experience some degree of alleviation in one or more characteristics of a disease or disorder; and/or some degree of alleviation in one or more symptoms caused by a disease or disorder, after receiving treatment with a therapeutic agent.
A “non-responder” as used herein, is a subject that will not experience some degree of alleviation in one or more characteristics of a disease or disorder after receiving treatment with a therapeutic agent; nor some degree of alleviation in one or more symptoms caused by a disease or disorder, after receiving treatment with the therapeutic agent.
A “reference criterion” as used herein, refers to a characteristic forming the basis of comparison for the evaluation or assessment of a measured characteristic.

Cancer and Cancer Stem Cells

Cancer is one of the most significant health conditions and leading causes of death worldwide. Currently available treatments include chemotherapy, radiation, surgery, hormonal therapy, immunotherapy, epigenetic therapy, anti-angiogenesis inhibitors, and other modalities, including targeted therapies, such as tyrosine kinase inhibitors and antibody based therapies. However, these treatments are ineffective in treating many cancers, and/or preventing reoccurrence. This ineffectiveness or unsustainability may be due, at least in part, to the innate heterogenic nature of cancer.
Cancers are known to be heterogeneous entities, with subsets of cancer cells exhibiting distinct molecular characteristics, including distinct gene expression profiles. Furthermore, cells with different molecular characteristics within the same cancer can respond differently to a single treatment. Cancer stem cells, cancer associated mesenchymal cells, and tumor initiating cancer cells, comprise a unique subpopulation of a tumor and have been identified in a large variety of cancer types. Relative to the remaining portion of the tumor, i.e., the tumor bulk, this subset of cancer cells is more tumorigenic, more slow growing or quiescent, and often more resistant to chemotherapeutic agents. Although, this subpopulation of cells constitutes only a small fraction of a tumor, these cells are thought to be responsible for cancer initiation, growth, and recurrence.
Given that currently available cancer treatments have, in large part, been designed to attack rapidly proliferating cells (i.e. those cancer cells that comprise the tumor bulk); cancer stem cells, cancer associated mesenchymal cells, and tumor initiating cancer cells, which are often slow growing, may be relatively more resistant to these treatments. Therefore, methods to identify cancer patients likely to respond positively to a treatment comprising an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells are needed; and can provide the basis for subsequent administration of a treatment comprising an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells to this candidate group of cancer patients.
The present invention provides a method of classifying subjects likely to respond to a particular therapeutic regimen for treating cancer. The method is based, at least in part, on the characterization of signals (e.g., the level of gene expression) possessed by a candidate subject population for treatment with a preselected drug. In general, the method involves identifying differences in candidate and non-candidate subject populations, where for example, a subject population has a gene expression profile associated with a candidate or non-candidate classification. The method can further include administration of the therapeutic regimen to the candidate population based on the characterized gene expression profile.
Overall, the invention described herein methods of evaluating and/or treating a subject, including acquiring a value or values that is a function of the level of gene expression for (each of) a plurality of genes selected from a first and/or second and/or third and/or fourth and/or fifth and/or sixth and/or or seventh and/or eighth set of genes; responsive to the value or values, classifying the subject as a candidate or non-candidate for treatment with a preselected drug; optionally, further treating the subject by administering the agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells; or withholding treatment from the subject; provided that if an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells is not administered, the acquisition of the subject sample or the acquisition of the value or values that is a function of the level of gene expression comprises directly acquiring; thereby evaluating or treating the subject. In response to the value or values, the invention also features: stratification of a subject population; identification or selection of the subject as likely or unlikely to respond positively to a treatment; selection of a treatment; or prognostication of the time course of the disease in the subject; measuring the response at the end of therapy and predicting the long term outcome; and/or determining the cancer stem cell population as a predictor of response to a treatment or therapy.

Subject Sample

The present invention features methods including, acquiring a subject sample. The terms “subject sample” and “sample” are used interchangeably herein. The subject sample can be a tissue, or bodily fluid, or bodily product. Tissue samples can include fixed, paraffin embedded, fresh, or frozen samples. For example, the tissue sample can include a biopsy, cheek swab, fine needle aspirates, large core needle biopsy, or directional vacuum assisted biopsy. Exemplary tissues include breast, brain, lung, pancreas, colon, prostate, lymph node, skin, hair follicles and nails. The tissue sample can also include a blood sample in which circulating tumor cells have been captured or isolated. Exemplary bodily fluids include blood, plasma, urine, lymph, tears, sweat, saliva, semen, and cerebrospinal fluid. Exemplary bodily products include exhaled breath.
The sample tissue, fluid, or product can be analyzed for the level of gene expression of a gene or a plurality of genes. The sample tissue, fluid or product can be analyzed for the level of gene expression of a gene or plurality of genes of a preselected signaling pathway or phenotypic pathway, e.g., a cancer stem cell phenotype, cancer associated mesenchymal cell phenotype, tumor initiating cancer cell phenotype, the epithelial to mesenchymal transition pathway, the Wnt signaling pathway, Notch pathway, or the TGFbeta signaling pathway. The sample tissue, fluid or product can be analyzed for the level of gene expression of a combination of genes from a plurality of preselected signaling or phenotypic pathways.
The tissue, fluid or product can be removed from the patient and analyzed. The evaluation can include one or more of: performing the analysis of the tissue, fluid or product; requesting analysis of the tissue fluid or product; requesting results from analysis of the tissue, fluid or product; or receiving the results from analysis of the tissue, fluid or product.
Acquisition of a Value or Values that is a Function of the Level of Gene Expression
The present invention features methods including, acquiring a value or values that is a function of the level of gene expression of a plurality of genes in a subject sample. The acquired value or values can be a function of a comparison with a reference criterion. The value or values can also be a function of the determination of whether the level of gene expression has a preselected relationship with a reference criterion (e.g., comparing the level of gene expression, with a preselected reference criterion). The reference criterion, as used herein, refers to a characteristic forming the basis of comparison for the evaluation or assessment of a measured characteristic. The preselected reference criterion can include the level of gene expression of a reference gene or the level of gene expression of a group of reference genes (e.g., housekeeping genes). The preselected reference criterion can include the level of gene expression of a gene from a control sample, e.g., a non-cancer sample. The appropriate reference criterion will depend on the gene or genes of which the level of expression is being acquired and the sample from which the level of gene expression of the genes was acquired from, and can be determined by one skilled in the art.
At least one or both of, acquiring a value or values that is the function of the level of gene expression, and determining if the level of gene expression has a preselected relationship with a reference criterion; can include one or more of: analyzing the sample, requesting analysis of the sample, requesting results from analysis of the sample, or receiving the results from analysis of the sample. Generally, analysis can include one or both of performing the underlying method (e.g., analysis of the level of gene expression) or receiving data from another who has performed the underlying method.
The acquired value or values can also be a function of a weighting factor. A weighting factor as used herein, refers to an element used to give an adjustment factor to a value. The weighting factor can be a composite weighting factor for a group of genes. For example, a first value or values that is a function of the level of gene expression of a plurality of genes can be a function of a weighting factor. The weighting factor can also be a specific weighting factor for a specific gene that only applies to that specific gene. For example, a first value or values that is a function of the level of gene expression of a first gene can be a function of a weighting factor, and a second value or values that is a function of the level of gene expression of a second gene can be a function of a second weighting factor; the first and the second weighting factor can be different.

Level of Gene Expression

The present invention features methods of acquiring a value or values that is a function of the level of gene expression of a plurality of genes. To acquire the level of gene expression in a subject sample, the level of gene expression can be assayed, such as by measuring the level of RNA or protein product produced by the relevant gene. Thus the level of gene expression can be a function of the level of a RNA product produced by the relevant gene; or the level of gene expression can be a function of the level of a protein product produced by the relevant gene. The level of gene expression can also be a function of the protein or RNA activity level, which can be assayed by determining the protein (or RNA, e.g., mRNA) activity levels, e.g., transcriptional activation activity, catalytic activity, gene silencing activity, kinase activity, etc. The level of RNA expression can be assayed by a PCR based method. For example, mRNA can be isolated from a tissue sample, and subjected to qRT-PCR, and, optionally, Southern blot analysis, or gene chip or microarray analysis or some variant thereof. The subject sample, or the mRNA isolated from, or amplified from, the subject sample, can be applied to a nucleic acid microarray, or chip array. The level of RNA expression can also be measured by, for example, RNA in situ hybridization, quantitative RNA sequencing, or Northern blot. The level of protein product expressed by the relevant gene can be assayed by various antibody based techniques, including but not limited to Western blot, immunohistochemistry, and immunoassays, e.g. ELISA. The levels of gene expression, e.g., level of RNA expression of the relevant gene, level of protein expression of the relevant gene; can be assayed by other molecular biology methods known to those skilled in the art.
Optionally, the level of gene expression data can be configured into a file, such as a data file, e.g., an image corresponding to the levels of gene expression. Optionally, the gene expression data can be stored in a tangible medium and/or transmitted to a second site. The evaluation of the data file or image can include one or more of, performing statistical data analysis or imaging analysis, requesting statistical data analysis or imaging analysis, requesting results from statistical data analysis or imaging analysis, or receiving the results from data statistical analysis or imaging analysis.

Location Specific Acquisition of the Level of Gene Expression

The present invention features methods which include the acquisition of a value or values for locations in the subject sample. The value or values can be a function of the level of gene expression of a gene or plurality of genes at the location. This can include the acquisition of a first value or values for a first location in the subject sample, and a second value or values for a second location in the subject sample, in which the value or values are a function of the level of gene expression of a gene or plurality of genes at the location. The term, “location”, as used herein, refers to a zone of a sample defined by preselected criteria, such as morphology, histopathology, and other attributes. A zone of a tumor can be defined by a unique gene expression pattern of a set of preselected genes. A zone may be classified as containing specific cell type or multiple cell types, e.g., a zone may be classified as a nodule of cancer stem cells, a nodule of cancer associated mesenchymal cells, a nodule of tumor initiating cancer cells; a zone of transition, e.g., an area between epithelial and mesenchymal features of a tumor region; or a boundary between tumor regions of different types; or it may be a niche indicated by the presence of a particular cell type or class, e.g., mesenchymal cells, stromal cells, inflammatory cells, endothelial cells, cancer stem cells, cancer associated mesenchymal cells, tumor initiating cancer cells, etc.
The level of gene expression at a location can be measured by RNA in situ hybridization and/or antibody based immunohistochemistry techniques. These techniques also allow for the association of the levels of gene expression with specific cell types in a zone or region through further definition or identification of the cells. The definition or identification of these cells can be assayed using computational overlays of the cells with specific gene markers of interest, or for adjoining cells. For example, an overlay may be achieved by evaluation of serial sections of formalin-fixed or frozen tumor tissues that are sectioned 3-5 microns in thickness. Adjoining sections may be evaluated with different probes, and computational methods applied to condense into a single image file with pseudocoloring representative of the different probes. Alternatively, probes that may be identified in different wavelength channels may be used together. The definition or identification of these cells can be determined by assaying the level of expression of gene markers of interest; or assaying the level of expression of gene markers of interest in adjoining cells. The definition or identification of the cells can also be assayed by histopathology criteria, e.g., cell shape, cell size, shape of cell, nucleus shape, nucleus size, and nuclei morphology, e.g., fuzzy nuclei.
The location in the subject sample can be defined, for example, as a distance from a morphological region of the subject sample, e.g., distance from an endothelial cell or blood vessel. The location can be the whole subject sample, e.g., a tumor sample. A first location can be the whole subject sample; with subsequent acquisition of the level of gene expression of a subset of genes that define a specific zone, e.g., zones defined by biological criteria, such as detection of genes associated with a specific identity, e.g., cancer stem cell, EMT, vasculature, etc.
The acquired value or values of each location can be a function of a comparison with a reference criterion. The value or values can be a function of the level of expression of a single gene at the location or a function of a combination of the level of gene expression of multiple genes at the location. For example, the level of gene expression of a group of genes can be measured with a uniform technique so that the collective expression of a set of genes together is acquired. For example, RNA in situ hybridization techniques can be used in which probe sets are used for two or more genes of interest that may be combined for analysis of subject samples.
The acquired value or values can be a function of a comparison with a reference criterion. The value or values can also be a function of the determination of whether the level of gene expression has a preselected relationship with a reference criterion (e.g., comparing the level of gene expression, with a preselected reference criterion). The reference criterion, as used herein, refers to a characteristic forming the basis of comparison for the evaluation or assessment of measured characteristic. The preselected reference criterion can include the level of gene expression of a reference gene or the level of gene expression of a group of reference genes (e.g., housekeeping genes). The preselected reference criterion can include the level of gene expression of a gene from a control sample, e.g., a non-cancer sample. The determination of whether the level of gene expression has a preselected relationship with a reference criterion can also include comparing the acquired value or values of a first location with the acquired value or values of a second location.
At least one or both of acquiring a value or values that is the function of the level of gene expression at a first and/or second location, and determining if the level of gene expression has a preselected relationship with a reference criterion, can include one or more of the following: analyzing the sample; requesting analysis of the sample; requesting results from analysis of the sample; or receiving the results from analysis of the sample. Generally, analysis can include one or both of performing the underlying method (e.g., analysis of the level of gene expression) or receiving data from another who has performed the underlying method.
The value or values of a first location can be associated with a higher or lower likelihood of being a cancer stem cell, cancer associated mesenchymal cell, or tumor initiating cancer cell, than a second value or values of a second location. The value or values of a first location can be associated with a higher or lower likelihood of being a cancer stem cell than a second value or values of a second location. The value or values of a first location can be associated with a higher or lower likelihood of being a cancer associated mesenchymal cell than a second value or values of a second location. The value or values of a first location can be associated with a higher or lower likelihood of being a tumor initiating cancer cell than a second value or values of a second location. Responsive to the acquisition of the value or values acquired for each of a plurality of locations, each location can be classified as being indicative of a cancer stem cell or non-cancer stem cell. For example, a location indicative of a cancer stem cell or a tumor initiating cancer cell can exhibit a high level of CD44 gene expression (CD44(high)) and a concurrent low level of CD24 gene expression (CD24(low)) compared to a reference criterion; an increased level of gene expression compared to a reference criterion of an EMT (epithelial to mesenchymal transition) transcription factor, e.g., ZEB1, Twist, FoxC2; a decreased level of gene expression compared to a reference criterion of tight junction and adhesion genes, e.g., Claudin1-7, E-cadherin; an increased level of gene expression of mesenchymal adhesion proteins, e.g., N-cadherin. Responsive to the acquisition of the value or values acquired for each of a plurality of locations, each location can be classified as a cancer stem cell or non-cancer stem cell. Each location can also be classified as a cancer stem cell, a cancer associated mesenchymal cell, or a tumor initiating cancer cell.
Where the value or values of a location are a function of the level of gene expression of multiple genes, the value or values can be indicative of a cancer stem cell, cancer associated mesenchymal cell, or tumor initiating cancer cell. For example, the level of gene expression of a set of genes can be measured with a uniform technique as described above so that the collective level of expression of the genes identify cancer stem cells, cancer associated mesenchymal cells, or tumor initiating cancer cells. Where the value or values of a location are a function of the level of gene expression of multiple genes, the value or values can be indicative of a cancer stem cell. For example, the level of gene expression of a set of genes can be measured with a uniform technique as described above so that the collective level of expression of the genes identifies cancer stem cells. Where the value or values of a location are a function of the level of gene expression of multiple genes, the value or values can be indicative of a cancer associated mesenchymal cell. For example, the level of gene expression of a set of genes can be measured with a uniform technique as described above so that the collective level of expression of the genes identifies cancer associated mesenchymal cells. Where the value or values of a location are a function of the level of gene expression of multiple genes, the value or values can be indicative of a tumor initiating cancer cell. For example, the level of gene expression of a set of genes can be measured with a uniform technique as described above so that the collective level of expression of the genes identifies tumor initiating cancer cells.
The locations can be separated by no distance, i.e., adjoining locations, in the subject sample or separated by range of distances; up to the maximum distance allowed by the sample size. For example, the locations can be separated by zero microns, ten microns, twenty microns, thirty microns, forty microns, fifty microns, sixty microns, seventy microns, eighty microns, ninety microns, one hundred microns, one hundred and fifty microns, two hundred microns, or three hundred microns; the locations can be separated by more than zero microns, more than ten microns, more than twenty microns, more than thirty microns, more than forty microns, more than fifty microns, more than sixty microns, more than seventy microns, more than eighty microns, more than ninety microns, more than one hundred microns, more than one hundred and fifty microns, more than two hundred microns, or more than three hundred microns; separated by at least one micron but not over one hundred microns; separated by at least fifty microns but not over one hundred microns; separated by at least one hundred microns; separated by at least one hundred microns but not more than two hundred microns; separated by at least two hundred microns but not more than three hundred microns; separated by at least three hundred microns; separated by at least four hundred microns; separated by at least five hundred microns; separated by at least six hundred microns, separated by at least seven hundred microns, separated by at least eight hundred microns, separated by at least nine hundred microns; separated by at least one thousand microns; separated by a distance over one thousand microns; separated by a distance under one thousand microns. The distance between locations can be any distance between zero and the maximum distance two locations can be separated based on the size of the sample, including zero and the maximum distance two locations can be separated based on the size of the sample.
The average distance between the locations can be zero microns; ten microns; twenty microns; thirty microns; forty micron; fifty microns; sixty microns; seventy microns; eighty microns; ninety microns; or one hundred microns. The average distance between the locations can be more than zero microns; more than ten microns; more than twenty microns; more than thirty microns; more than forty micron; more than fifty microns; more than sixty microns; more than seventy microns; more than eighty microns; more than ninety microns; or more than one hundred microns. The average distance between the locations can be more than one thousand microns. The average distance between the locations can be more than one hundred microns; more than 200 hundred microns; more than three hundred microns; more than four hundred microns; more than five hundred microns, or more than one thousand microns. The average distance between locations can be any distance between zero and the maximum distance two locations can be separated based on the size of the sample, including zero and the maximum distance two locations can be separated based on the size of the sample.

Gene Set Score

The present invention features methods of acquiring a gene set score. The gene set score can be a function of the level of gene expression of a plurality of genes. The level of gene expression can be acquired as described above. The gene set score can further be a function of the level expression of a gene isoform. The level of a gene isoform can be acquired as described above. The gene set score can be a function of both the level of gene expression and the level of expression of a gene isoform. The gene set score can be a function of both the level of gene expression and the level of expression of a plurality of gene isoforms of a gene. The gene set score can be a function of both the level of gene expression of a gene or plurality of genes; and the level of expression of a gene isoform of a gene. The gene set score can be a function of the level of gene expression of a gene or plurality of genes; and the level of expression of each gene isoform of a plurality of gene isoforms of a gene. The gene set score can be a function of both the level of gene expression of a gene or plurality of genes; and the level of expression of a plurality of gene isoforms of a gene. The set gene score can be a function of both the level of gene expression of a gene or plurality of genes; and the level of expression of a plurality of gene isoforms of a plurality of genes. The gene set score can be a function of both the level of gene expression of a gene or plurality of genes; and the level of expression of each gene isoform of a plurality of gene isoforms of a plurality of genes.
The gene set score can be acquired by mathematical computation. The gene set score can be computed using the following algorithm:
$S_{sig_X} = \frac{1}{N} \sum_{i = 1}^{N} (e_{i} - {\overline{e}}_{i})$

Where:

S_sig _— _X=the score for a subset of the genes in the signature gene set (i.e., S_sig _— _UPor S_sig _— _DN)
N=number of genes in the gene set
e_i=the log 2 expression level of gene i in the gene set
ē_i=the mean log 2 expression level of gene i over all samples in the sample set Gene set score:
S _sig =S _sig _— _UP −S _sig _— _DN

Where:

S_sig _— _UP=gene set score over upregulated genes in the signature
S_sig _— _DN=gene set score over downregulated genes in the signature.

Genotype

The present invention features methods that include the acquisition of a genotype of the subject sample. The subject sample can be any sample type described herein, e.g., a tissue sample, bodily fluid, or bodily product. The genotype can be directly acquired or indirectly acquired. The genotype can be directly acquired through assaying. The genotype can be assayed using a sequencing based method. “Sequencing” a nucleic acid molecule as used herein, requires determining the identity of at least one nucleotide in the molecule. The identity of less than all of the nucleotides in a molecule can be determined. The identity of a majority or all of the nucleotides in the molecule can be determined. The genotype can be assayed using a sequencing based method, e.g., SNP (single nucleotide polymorphism) analysis, PCR based method, restriction fragment length polymorphism, terminal restriction fragment length polymorphism, amplified restriction fragment length polymorphism, multiplex restriction fragment length polymorphism, or other sequencing and molecular biology techniques known to those skilled in the art.
In genotyping, genetic events associated with cancer can be assayed. For example, nucleotides of the sample can be sequenced to determine the presence or absence of a genetic event associated with cancer; an oncogene or oncogenes and/or tumor suppressor genes can be sequenced, e.g., Abl, Af4/hrx, akt-2, alk, alk/npm, aml 1, aml 1/mtg8, APC, axl, bcl-2, bcl-3, bcl-6, bcr/abl, brca-1, brca-2, beta-catenin, CDKN2, c-myc, c-sis, dbl, dek/can, E2A/pbx1, egfr, enl/hrx, erg/TLS, erbB, erbB-2, erk, ets-1, ews/fli-1, fms, fos, fps, gli, gsp, HER2/neu, hox11, hst, IL-3, int-2, jun, kit, KS3, K-sam, Lbc, lck, lmo1, lmo2, L-myc, li1-1, lyt-10, lyt-10/C alpha1, mas, mdm-2, mll, mos, mtg8/aml1, myb, myc, MYH11/CBFB, neu, nm23, N-myc, ost, p53, pax-5, pbx1/E2A, pdgfr, PI3-K, pim-1, PRAD-1, raf, RAR/PML, rash, rasK, rasN, Rb, rel/nrg, ret, rhom1, rhom2, ros, ski, sis, set/can, src, tal1, tal2, tan-1, telomerase, Tiam1, TSC2, trk, vegfr, or wnt.

Classification

The present invention features methods including, classifying the subject, e.g., classifying the subject as a candidate or a non-candidate for treatment with a preselected drug, e.g., an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells. As used herein, a subject who is a “candidate” is a one more likely to respond to a particular therapeutic regimen, relative to a reference subject or group of subjects. A “non-candidate” subject is one not more likely to respond to a particular therapeutic regimen, relative to a reference subject or group of subjects. The preselected drug can include but is not limited to, an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells; which can include but is not limited to, e.g., salinomycin; a gamma secretase inhibitor; a DLL4 inhibitor, e.g., a therapeutic antibody targeting DLL4; a TRAIL inhibitor, e.g., a therapeutic antibody targeting TRAIL; a Hedgehog inhibitor, e.g., a therapeutic antibody targeting Hedgehog; a NOTCH3 inhibitor, e.g., a therapeutic antibody targeting NOTCH3; a NOTCH4 inhibitor, e.g., a therapeutic antibody targeting NOTCH4; a panNOTCH inhibitor, e.g., a therapeutic antibody targeting panNOTCH; a FGFR1 inhibitor, e.g., a therapeutic antibody targeting FGR1; a FGFR2 inhibitor, e.g., a therapeutic antibody targeting FGR2; a FGFR3 inhibitor, e.g., a therapeutic antibody targeting FGR3; a FGFR4 inhibitor, e.g., a therapeutic antibody targeting FGR4; a RON inhibitor, e.g., a therapeutic antibody targeting RON; Wnt pathway inhibitor, e.g., therapeutic antibodies targeting the Wnt pathway; a PI3Kinase inhibitor; a mTOR inhibitor; sodium meta arsenite; verapail; reserpine; a perifosen inhibitor of FAK1; a FAK inhibitor; a p38 inhibitor. Classification as a candidate subject can also reflect an increased likelihood the subject will respond positively to treatment with an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells.

Administration

The present invention features methods including, administering a treatment comprising an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells to the subject. The invention can further include selecting a regimen, e.g., dosage, formulation, route of administration, number of dosages, or adjunctive or combination therapies of an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells. The administration of an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells can be responsive to the acquisition of the value or values that is a function of the level of gene expression described herein, and/or classification of a subject as a candidate or non-candidate for treatment with an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells. The selection of the regimen can be responsive to the acquisition of the value or values that is a function of the level of gene expression described herein, and/or classification of a subject as a candidate or non-candidate for treatment with an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells. The invention can further include the administration of the selected regimen. The administration can be provided responsive to acquiring knowledge or information of the value or values that is a function of the level of gene expression described herein, from another party; receiving communication of the presence of the value or values that is a function of the level of gene expression in a subject; or responsive to the acquisition of the value or values that is a function of the level of gene expression in a subject, wherein the acquisition arises from collaboration with another party.
An agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells, e.g., salinomycin; a gamma secretase inhibitor; a DLL4 inhibitor, e.g., a therapeutic antibody targeting DLL4; a TRAIL inhibitor, e.g., a therapeutic antibody targeting TRAIL; a Hedgehog inhibitor, e.g., a therapeutic antibody targeting Hedgehog; a NOTCH3 inhibitor, e.g., a therapeutic antibody targeting NOTCH3; a NOTCH4 inhibitor, e.g., a therapeutic antibody targeting NOTCH4; a panNOTCH inhibitor, e.g., a therapeutic antibody targeting panNOTCH; a FGFR1 inhibitor, e.g., a therapeutic antibody targeting FGR1; a FGFR2 inhibitor, e.g., a therapeutic antibody targeting FGR2; a FGFR3 inhibitor, e.g., a therapeutic antibody targeting FGR3; a FGFR4 inhibitor, e.g., a therapeutic antibody targeting FGR4; a RON inhibitor, e.g., a therapeutic antibody targeting RON; Wnt pathway inhibitor, e.g., therapeutic antibodies targeting the Wnt pathway; a PI3Kinase inhibitor; a mTOR inhibitor; sodium meta arsenite; verapail; reserpine; a perifosen inhibitor of FAK1; a FAK inhibitor; a p38 inhibitor; can be administered to a subject using any amount and any route of administration effective for treating cancer, or symptoms associated with cancer. The exact dosage required will vary from subject to subject, depending on subject specific factors, e.g., the age and general condition of the subject, concurrent treatments, concurrent diseases or conditions; cancer specific factors, e.g., the type of cancer, whether the cancer is recurrent, whether the cancer is metastatic, the severity of the disease; and agent specific factors., e.g., its composition, its mode of administration, its mode of activity, and the like. For example, the dosage may vary depending on whether the subject is currently receiving or had previously received a treatment regimen prior to the administration of an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells; whether the subject is a non-responder to such current or previous treatment; whether the subject's cancer is recurrent; or whether the subject's cancer has metastasized to a second tissue site.
The total daily usage of a therapeutic composition of an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells can be decided by an attending physician within the scope of sound medical judgment. The specific therapeutically effective, dose level for any particular subject will depend upon a variety of factors including the type of cancer being treated; the severity of the cancer; the metastatic state of the cancer; the recurrence state of the cancer; the activity of the specific compound employed; the specific composition employed; the age, body weight, general health, sex and diet of the patient; the time of administration, route of administration, and rate of excretion of the specific compound employed; the duration of the treatment; drugs used in combination or coincidental with the specific compound employed; and like factors well known in the medical arts.
The agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells may be administered by any route, including by those routes currently accepted and approved for known products. Exemplary routes of administration include, e.g., oral, intraventricular, transdermal, rectal, intravaginal, topical (e.g. by powders, ointments, creams, gels, lotions, and/or drops), mucosal, nasal, buccal, enteral, vitreal, sublingual; by intratracheal instillation, bronchial instillation, and/or inhalation; as an oral spray, nasal spray, and/or aerosol, and/or through a portal vein catheter. An agent may be administered in a way, which allows the agent to cross the blood-brain barrier, vascular barrier, or other epithelial barrier.
Other exemplary routes include administration by a parenteral mode (e.g., intravenous, subcutaneous, intraperitoneal, or intramuscular injection). The phrases “parenteral administration” and “administered parenterally” as used herein mean modes of administration other than enteral and topical administration, usually by injection, and include, without limitation, intravenous, intramuscular, intraarterial, intrathecal, intracapsular, intramedullary, intratumoral, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, subcuticular, intraarticular, subcapsular, subarachnoid, intraspinal, epidural and intrasternal injection and infusion.
Pharmaceutical compositions can be formulated in a variety of different forms, such as liquid, semi-solid and solid dosage forms, such as liquid solutions (e.g., injectable and infusible solutions), dispersions or suspensions, tablets, pills, powders, liposomes and suppositories. The preferred form can depend on the intended mode of administration and therapeutic application. A pharmaceutical composition comprising an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells may be administered on various dosing schedules. The dosing schedule will be dependent on several factors including, the type of cancer being treated; the severity of the cancer; the metastatic state of the cancer; the recurrence state of the cancer; the activity of the specific compound employed; the specific composition employed; the age, body weight, general health, sex and diet of the patient; the time of administration, route of administration, and rate of excretion of the specific compound employed; the duration of the treatment; drugs used in combination or coincidental with the specific compound employed; and like factors well known in the medical arts.
Exemplary dosing schedules of an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells composition include, once daily, or once weekly, or once monthly, or once every other month. The composition can be administered twice per week or twice per month, or once every two, three or four weeks. The composition can be administered as two, three, or more sub-doses at appropriate intervals throughout the day or even using continuous infusion or delivery through a controlled release formulation. In that case, the therapeutic agent contained in each sub-dose may be correspondingly smaller in order to achieve the total daily dosage. The dosage can also be compounded for delivery over several days, e.g., using a conventional sustained release formulation, which provides sustained release of the agent over a several day period. Sustained release formulations are well known in the art and are particularly useful for delivery of agents at a particular site.
The present invention features methods in which a value or values that is a function of the level of gene expression for (each of) a plurality of genes can be acquired at the time of or after diagnosis of cancer in a subject. The acquisition of the value or values that is a function of the level of gene expression can be acquired at a predetermined interval, e.g., a first point in time and at least at a subsequent point in time. The cancer can include cancers characterized as comprising cancer stem cells, cancer associated mesenchymal cells, or tumor initiating cancer cells. The cancer can include cancers that have been characterized as being enriched with cancer stem cells, cancer associated mesenchymal cells, or tumor initiating cancer cells. Exemplary cancers include epithelial cancers, breast, lung, pancreatic, colorectal, prostate, head and neck, melanoma, acute myelogenous leukemia, and glioblastoma. Exemplary breast cancers include triple negative breast cancer, basal-like breast cancer, claudin-low breast cancer, invasive, inflammatory, metaplastic, and advanced Her-2 positive or ER-positive cancers resistant to therapy. Other cancers include but are not limited to, brain, abdominal, esophagus, gastrointestinal, glioma, liver, tongue, neuroblastoma, osteosarcoma, ovarian, retinoblastoma, Wilm's tumor, multiple myeloma, skin, lymphoma, blood, retinal, acute lymphoblastic leukemia, bladder, cervical, kidney, endometrial, meningioma, lymphoma, skin, uterine, lung, non small cell lung, nasopharyngeal carcinoma, neuroblastoma, solid tumor, hematologic malignancy, leukemia, squamous cell carcinoma, testicular, thyroid, mesothelioma, brain vulval, sarcoma, intestine, oral, T cell leukemia, endocrine, salivary, spermatocytic seminoma, sporadic medulalry thyroid carcinoma, non-proliferating testes cells, cancers related to malignant mast cells, non-Hodgkin's lymphoma, and diffuse large B cell lymphoma.
The cancer can be a primary tumor, i.e., located at the anatomical site of tumor growth initiation. The cancer can also be metastatic, i.e., appearing at least a second anatomical site other than the anatomical site of tumor growth initiation. The cancer can be a recurrent cancer, i.e., cancer that returns following treatment, and after a period of time in which the cancer was undetectable. The recurrent cancer can be anatomically located locally to the original tumor, e.g., anatomically near the original tumor; regionally to the original tumor, e.g., in a lymph node located near the original tumor; or distantly to the original tumor, e.g., anatomically in a region remote from the original tumor.
The acquisition of a value or values that is a function of the level of gene expression described herein, can be acquired prior to, during, or after administration of a treatment to a subject. The treatment can include an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells therapy. The treatment can include a chemotherapeutic agent, antiemetic, analgesic, or anti-inflammatory agent. Suitable chemotherapeutic agents are any chemical substances or compounds, such as cytotoxic or cytostatic agent, that is used to treat a condition, particularly cancer, including, but not limited to: alkylating agents (e.g., nitrogen mustards such as chlorambucil, cyclophosphamide, isofamide, mechlorethamine, melphalan, and uracil mustard; aziridines such as thiotepa; methanesulphonate esters such as busulfan; nitroso ureas such as carmustine, lomustine, and streptozocin; platinum complexes such as cisplatin and carboplatin; bioreductive alkylators such as mitomycin, procarbazine, dacarbazine and altretamine); DNA strand-breakage agents (e.g., bleomycin); topoisomerase II inhibitors (e.g., amsacrine, dactinomycin, daunorubicin, idarubicin, mitoxantrone, doxorubicin, etoposide, and teniposide); DNA minor groove binding agents (e.g., plicamydin); antimetabolites (e.g., folate antagonists such as methotrexate and trimetrexate; pyrimidine antagonists such as fluorouracil, fluorodeoxyuridine, CB3717, azacitidine, cytarabine, and floxuridine; purine antagonists such as mercaptopurine, 6-thioguanine, fludarabine, pentostatin; asparginase; and ribonucleotide reductase inhibitors such as hydroxyurea); tubulin interactive agents (e.g., vincristine, vinblastine, and paclitaxel (Taxol)); hormonal agents (e.g., estrogens; conjugated estrogens; ethinyl estradiol; diethylstilbesterol; chlortrianisen; idenestrol; progestins such as hydroxyprogesterone caproate, medroxyprogesterone, and megestrol; and androgens such as testosterone, testosterone propionate, fluoxymesterone, and methyltestosterone); adrenal corticosteroids (e.g., prednisone, dexamethasone, methylprednisolone, and prednisolone); leutinizing hormone releasing agents or gonadotropin-releasing hormone antagonists (e.g., leuprolide acetate and goserelin acetate); and antihormonal antigens (e.g., tamoxifen, antiandrogen agents such as flutamide; and antiadrenal agents such as mitotane and aminoglutethimide). Exemplary chemotherapeutic agents include, Capecitabine, Carboplatin, Cisplatin, Cyclophosphamide, Docetaxel, Doxorubicin, Epirubicin, Eribulin, mesylate5-Fluorouracil, Gemcitabine, Ixabepilone, Liposomal doxorubicin, Methotrexate, Paclitaxel, or Vinorelbine; or any combination thereof.
The subject can be a responder or non-responder to the current or prior treatment. The agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells; can be administered as an additional therapeutic agent, e.g., an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells in addition to a current therapeutic regimen, or in addition to a new therapeutic regimen. The current treatment of the subject can be stopped and replaced with treatment an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells. The current treatment regimen can also be altered with the addition of an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells as an additional therapeutic agent. Therapeutic agents administered in combination with an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells; can kill or inhibit the growth of non-cancer stem cells, non-cancer associated mesenchymal cells, or non-tumor initiating cells in the subject.

Kits or Products

The present invention features a kit or product that includes a means to assay the level of gene expression of a gene or plurality of genes in Table 1. For example, the kit or product can include an agent capable of interacting with a gene expression product of a gene from the genes in Table 1; and can further contain a second agent capable of interacting with a different gene expression product from a gene in Table 1. The kit can contain a plurality of different agents capable of interacting with a plurality of genes expression products from a gene in Table 1. The kit can contain a plurality of different agents capable of interacting with a plurality of genes expression products from a plurality of genes in Table 1. The agent can include, but is not limited to, an antibody, a plurality of antibodies, an oligonucleotide, or a plurality of oligonucleotides. The kit or product can further comprise an agent capable of interacting with a gene expression product of a gene not in Table 1. The kit or product can contain a plurality of agents capable of interacting with a plurality of gene expression product of a plurality of genes not in Table 1. The gene expression product can include, but is not limited to, a RNA product of the associated gene, or a protein product of the associated gene.
The kit or product can further optionally include reagents for performing the level of gene expression assays described herein. For example, the kit can include buffers, solvents, stabilizers, preservatives, purification columns, detection reagents, and enzymes, which may be necessary for isolating nucleic acids from a subject sample, amplifying the samples, e.g., by qRT-PCR, and applying the samples to the agent described above; or for isolating proteins from a subject sample, and applying the samples to the agent described above; or reagents for directly applying the subject sample to the agent described above. A kit can also include positive and negative control samples, e.g., control nucleic acid samples (e.g., nucleic acid sample from a non-cancer subject, or a non-tumor tissue sample, or a subject who has not received treatment for cancer, or other test samples for testing at the same time as subject samples. A kit can also include instructional material, which may provide guidance for collecting and processing patient samples, applying the samples to the level of gene expression assay, and for interpreting assay results.
The components of the kit can be provided in any form, e.g., liquid, dried, semi-dried, or in lyophilized form, or in a form for storage in a frozen condition. Typically, the components of the kit are provided in a form that is sterile. When reagents are provided in a liquid solution, the liquid solution generally is an aqueous solution, e.g., a sterile aqueous solution. When reagents are provided in a dried form, reconstitution generally is accomplished by the addition of a suitable solvent. The solvent, e.g., sterile buffer, can optionally be provided in the kit.
The kit can include one or more containers for the kit components in a concentration suitable for use in the level of gene expression assays or with instructions for dilution for use in the assay. The kit can contain separate containers, dividers or compartments for the assay components, and the informational material. For example, the positive and negative control samples can be contained in a bottle or vial, the clinically compatible classifier can be sealed in a sterile plastic wrapping, and the informational material can be contained in a plastic sleeve or packet. The kit can include a plurality (e.g., a pack) of individual containers, each containing one or more unit forms (e.g., for use with one assay) of an agent. The containers of the kits can be air tight and/or waterproof. The container can be labeled for use.
The kit can include informational material for performing and interpreting the assay. The kit can also provide guidance as to where to report the results of the assay, e.g., to a treatment center or healthcare provider. The kit can include forms for reporting the results of a gene activity assay described herein, and address and contact information regarding where to send such forms or other related information; or a URL (Uniform Resource Locator) address for reporting the results in an online database or an online application (e.g., an app). In another embodiment, the informational material can include guidance regarding whether a patient should receive treatment with an ant-cancer stem cell agent, depending on the results of the assay.
The informational material of the kits is not limited in its form. In many cases, the informational material, e.g., instructions, is provided in printed matter, e.g., a printed text, drawing, and/or photograph, e.g., a label or printed sheet. However, the informational material can also be provided in other formats, such as computer readable material, video recording, or audio recording. The informational material of the kit can be contact information, e.g., a physical address, email address, website, or telephone number, where a user of the kit can obtain substantive information about the gene activity assay and/or its use in the methods described herein. The informational material can also be provided in any combination of formats.
A subject sample can be provided to an assay provider, e.g., a service provider (such as a third party facility) or a healthcare provider that evaluates the sample in an assay and provides a read out. For example, an assay provider can receive a sample from a subject, such as a tissue sample, or a plasma, blood or serum sample, and evaluate the sample using an assay described herein, and determines that the subject is a candidate to receive an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells.
The assay provider can inform a healthcare provider that the subject is a candidate for treatment with an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells, and the candidate is administered the agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells. The assay provider can provide the results of the evaluation, and optionally, conclusions regarding one or more of diagnosis, prognosis, or appropriate therapy options to, for example, a healthcare provider, or patient, or an insurance company, in any suitable format, such as by mail or electronically, or through an online database. The information collected and provided by the assay provider can be stored in a database.

Reports

The present invention features optionally providing a report. The report can include a prediction of the likelihood that a subject will respond positively or will not respond positively to treatment with an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells, e.g., salinomycin; a gamma secretase inhibitor; a DLL4 inhibitor, e.g., a therapeutic antibody targeting DLL4; a TRAIL inhibitor, e.g., a therapeutic antibody targeting TRAIL; a Hedgehog inhibitor, e.g., a therapeutic antibody targeting Hedgehog; a NOTCH3 inhibitor, e.g., a therapeutic antibody targeting NOTCH3; a NOTCH4 inhibitor, e.g., a therapeutic antibody targeting NOTCH4; a panNOTCH inhibitor, e.g., a therapeutic antibody targeting panNOTCH; a FGFR1 inhibitor, e.g., a therapeutic antibody targeting FGR1; a FGFR2 inhibitor, e.g., a therapeutic antibody targeting FGR2; a FGFR3 inhibitor, e.g., a therapeutic antibody targeting FGR3; a FGFR4 inhibitor, e.g., a therapeutic antibody targeting FGR4; a RON inhibitor, e.g., a therapeutic antibody targeting RON; Wnt pathway inhibitor, e.g., therapeutic antibodies targeting the Wnt pathway; a PI3Kinase inhibitor; a mTOR inhibitor; sodium meta arsenite; verapail; reserpine; a perifosen inhibitor of FAK1; a FAK inhibitor; a p38 inhibitor. The report can include a prediction of the likelihood a subject will respond positively or not to treatment with an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells. The report can also include a proposal including any one of or combination of the following: whether a subject is a candidate for treatment with an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells; whether a subject should be treated with a preselected drug, e.g. an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells; or whether treatment with a preselected drug, e.g., an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells, should be withheld.
The report can be provided by an assay service provider (such as a third party facility) that evaluates the sample in an assay and provides a report, or a healthcare provider. In the former case, the assay service provider can inform a healthcare provider that the subject is a candidate for treatment with an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells, and the candidate is administered the agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells. The assay provider can provide the results of the evaluation, and optionally, conclusions regarding one or more of diagnosis, prognosis, or appropriate therapy options to, for example, a healthcare provider, or subject, or an insurance company, in any suitable format, such as by mail or electronically, or through an online database. The information collected and provided by the assay provider can be stored in a database. The report can be reported back to the healthcare provider, such as through a form, which can be submitted by mail or electronically (e.g., through facsimile or e-mail) or through an on-line database or on-line application (e.g., through an “app”). The results of the assay (including the level of gene expression) can be stored in a database and can be accessed by a healthcare provider, such as through the worldwide web.

EXAMPLES

Example 1

Set of Genes Derived from Cancer Stem Cell and EMT Identifiers Found in More than One Statistical Measure for Gene Ranking

Three statistical processes of gene sampling methods were utilized to rank genes that contribute to tumor initiating cancer cells, cancer stem cells, and cancer associated mesenchymal cells (FIG. 1). The three partitioning and ranking methods used were LASSO [LASSO Variable Selection; Tibshirani, R. (1996)], Recursive Partitioning (or CART), and Stepwise selection. Patient samples used to train the models were classified as likely to recur using an arbitrary threshold as follows, Rscore <0.7 as ‘predicted recurrence’. In the LASSO method, the process is to fit outcomes using all genes, with the added restriction that the sum of model parameters must be less than an arbitrary threshold. While increasing the threshold in small steps and re-fitting the outcomes model; the determined gene ranking is the order that the genes model parameter exceeds threshold. In the Stepwise method, the process is to fit outcomes using all genes, and successively dropping the least significant gene from the model, and re-fitting the outcomes model until model performance does not improve. AIC criterion is used as a measure of fit, which balances the number of parameters against the improvement in performance. In the CART method, the process is to successively choose from the list of all genes, a single gene that splits samples into groups with the smallest impurity of outcomes. This process splits each subgroup using a single gene, until the minimum group size is reached, or the impurity of outcomes cannot be improved.
In the LASSO gene ranking method, there is a selection of the first 20 genes to enter a model using a 1e−10 threshold, and opening up using regression status. The next step involves fitting a logit model using all 20 genes, and repeating a logit fit using the top 10 genes, and a logit fit using restriction to increased statistical significance in additional cycles.
In the STEPWISE gene ranking method, sixty (60) genes are chosen randomly from a gene list. A model is fit that predicts outcome with all 60 genes using all samples. The least significant genes (highest AIC) are dropped from the set and refit until predictive ability of the model is affected (AIC increases). The final list of genes is kept and their statistical significance in the final model (p-values) is determined. The procedure is repeated 100 times to capture variety of starting genes with 60 ‘different’ or random genes. The ‘best’ genes are chosen based on average significance level within all models.
In the CART gene ranking method, 5000 iterations are performed, sampling from a patient dataset to be evaluated, that is equally split between patients having had recurrences or non-recurrences. Random sets of 20 genes from the analysis group are selected and fit to outcome in a decision tree composed of 5 samples per leaf. Trimming is undertaken to reveal the most important branches (cp=0.01). Between 1 and 5 genes are kept for next step (median of 3 genes). In the counting strategy, the more often a gene is included in a model is indicative of a higher statistical importance of that gene.
A group of 52 genes were identified that were highly ranked by two of the three methods. A concordance of the genes from these three methods was observed, where 16 genes were found by all of the methods (FIG. 1 and FIG. 2), such as the genes: CYBRD1, ARTN, CITED2, SLC7A5, CHN1, CTGF, CSTA, MYL9, EPB41L4B, KRT15, LRIG1, CDH3, TPD52L1, ANXA3, OLFML3, and CORO1A.
Triple negative breast cancer patients (n=178) were evaluated by standard histopathology criteria and determined to be of the triple negative breast cancer subtype by examining ER, PR, and HER2 by immunohistochemistry and pathology of tumor biopsy specimen. Following their diagnosis, patients were treated with standard chemotherapy and their survival was evaluated over the next seven years, so that outcome data was available for the patient cohort. In one test, gene subsets identified by the above rank ordering strategies were evaluated in breast cancer patient specimens with RNA expression data formed from profiling of patient tumors. In addition, recurrence and/or survival data from the patients was utilized.
Using the top 20 genes of the LASSO method, namely genes: CYBRD1, ARTN, KRT15, ITGB4, RGS4, SPRR1B, SLC7A5, ANXA3, IL18, AP1M2, CHN1, OLFML3, ZBED2, LEPREL1, ABLIM1, CDH3, DNAJB4, TRAM2, UCHL1, and CTGF; analysis of patient and RNA data for these genes in Kaplan-Meier recurrence plots showed, the Event Free Recurrence subgroup of patients was significantly distinguished from the patient subset suffering recurrence by this 20-gene test (FIG. 3). In addition, the 20-gene model had a significant True Positive Rate over a False Positive Rate as exemplified by Area-Under the Curve (AUC) statistical analysis, with an AUC=0.88 (FIG. 3). The p-value separating the Event Free Recurrence subgroup from patients with Recurrence was determined to be p=8.88e−16. The top five ranked genes by this method displayed a separation of Even Free Recurrences with an AUC=0.73 and p=0.000423. Permutation analysis on LASSO method in this exercise was evaluated with a p=0.01 for 99.1 percentile for the top 20 gene model with regard to calculated p values, and a p=0.003 for 99.7 percentile for the top 20 gene model with regard to calculated AUCs.
Using the top 20 genes of the STEPWISE method, namely genes: CYBRD1, ARTN, RGS4, CITED2, SLC7A5, ITGB4, CHN1, PI3, TMEM40, NDRG1, SPRR1A, PCDH9, CTGF, MAP1B, DSG3, AP1M2, FBLN5, SERPINB13, PMP22, and CSTA; analysis of patient and RNA data for these genes in Kaplan-Meier recurrence plots showed, the Event Free Recurrence subgroup of patients was significantly distinguished from the patient subset suffering recurrence by this 20-gene test, with a highly significant p-value distinguishing the groups of p=2.01e−13 (FIG. 4). In addition, the 20-gene model had a significant True Positive Rate over a False Positive Rate as exemplified by Area-Under the Curve (AUC) statistical analysis, with an AUC=0.87 (FIG. 4).
Using the top 20 genes of the CART method, namely genes: CYBRD1, RGS4, CDH3, CHN1, SLC7A5, PRRX1, CITED2, COL5A2, ARTN, ZBED2, SNED1, ITGB4, LUM, PVRL3, OLFML3, BIN1, CCND2, DAB2, ANXA3, and IL18; analysis of patient and RNA data for these genes in Kaplan-Meier recurrence plots showed, the Event Free Recurrence subgroup of patients was significantly distinguished from the patient subset suffering recurrence by this 20-gene test (FIG. 5). In addition, the 20-gene model has a significant True Positive Rate over a False Positive Rate as exemplified by Area-Under the Curve (AUC) statistical analysis, with an AUC=0.93 (FIG. 5).
Utilizing these methods, gene sets were determined that identify recurrence-free patient subgroups from Kaplan-Meier analysis. Importantly, using genes that identify cancer stem cells and the epithelial-mesenchymal transition, associated with their gene expression, key models were discovered that may be utilized in a diagnostic platform for patient discrimination. As this example is displayed for breast cancer, and particularly for triple negative breast cancer, the method is of importance in these cancers. In all likelihood, additional value may be extended to predicting outcome for other cancer types, particularly other solid tumor types.

Example 2

Determination of Candidate Cancer Stem Cell Genes that Increase the Performance of Gene Set Models for Identifying the Recurrence-Free Patient Subset in Breast Cancer

In another example, genes that significantly contribute to the ability of gene expression signatures to predict recurrence of cancer were discovered by a process of combining genes with other gene sets. In this exercise, a similar evaluation of breast cancers was explored as an example of the method of discovery and identification of important genes. These genes may describe properties of cancer stem cells, tumor initiating cancer cells, cancer stem cells, and cancer associated mesenchymal cells. This example was conducted on triple negative breast cancer, but it is likely not to be limited to this cancer type or disease. In the present example triple negative breast cancer patients (n=178) were evaluated by standard histopathology criteria and determined to be of the triple negative breast cancer subtype by examining ER, PR, and HER2 by immunohistochemistry and pathology of tumor biopsy specimen. Following their diagnosis, patients were treated with standard chemotherapy and their survival was evaluated over the next seven years, so that outcome data was available for the patient cohort. In one test, gene subsets identified by the above described rank ordering strategies were evaluated in breast cancer patient specimens with RNA expression data formed from profiling patient tumors. In addition, recurrence and/or survival data from the patients was utilized.
Genes used in the addition test were at least one from the group of: AXL, NOTCH3, GPR30, PROCR, PAX2, PI3KA, CD44, CD133, ALDH1, BMI1, KRT19, NRP1, MSI1, MTOR, JAG1, p38, ESA1, ZEB1, TWIST1, FOXC2, CDH2, ZEB2, FOXC1, SNAI1, SNAI2, TWIST2, GSC, TCF3, CD44, VIM, FN1, MLPH1, and ARHGEF11. In the procedure to discover the significance of these genes towards a recurrence prediction, the following test was executed. First, a set of genes was selected from the gene list (i); also shown in the Table 1. In this example, the base model from this group was identified by the LASSO procedure. Eight of the genes that were moderately ranked [Rank 6-13] were selected [SPRR1B, SLC7A5, ANXA3, IL18, AP1M2, CHN1, OLFML3, and ZBED2] after trimming of the top five genes on the LASSO output list. In the analysis, gene addition benefit is demonstrated by comparison of a ‘positive control’ gene dictated by the highest ranking genes of the LASSO set. In this analysis, any one of the following genes was used [CYBRD1, ARTN, KRT15, ITGB4, RGS4; Rank order 1-5]. To evaluate the importance of a new gene, a model was assembled with the Added gene or with a Positive Control gene. Both 5-gene models were then assessed to predict recurrence/non-recurrence patient subgroup identification with genes (logit model, 0.5 probability threshold for assignment). The method used the difference in recurrence curves of predicted classification as a performance measure.
As illustrated in FIG. 7 and FIG. 8, a number of candidate genes were identified to improve the statistical values of base models. These improvements were computed according to each of the genes designated in FIG. 7 and FIG. 8. The p-values of the addition of the candidate genes ranged from p=2.45e−8 to p=7.38e−4. For purposes of comparison, the p-values of addition of the Positive Control Gene ranged from p=4.23e−10 to p=4.70e−04. It is noteworthy that a very high percentage of cancer stem cell/epithelial-mesenchymal transition genes were shown to improve 5-gene models containing the new gene. According to the output of this method, important genes are interrogated for significance when combined in 5-gene models from the base 4-gene models. The method demonstrates the importance of the added genes. Gene models containing the new gene are illustrated to identify recurrence subgroups of triple negative breast cancer patients.
In addition to demonstrating individual contributions of driver genes, the same genes were evaluated as a group in Kaplan-Meier recurrence plots (FIG. 9). In one example, the genes of the candidate CSC genes [NOTCH3, PROCR, BMI1, KRT19, MSI1, NRP1, JAG1, ALDH1A1, and HTATIP] were used in a multi-gene model and in a Kaplan-Meier recurrence plot. The 9-gene model gave separation of triple negative breast cancer patients into the Recurrence and No Recurrence groups with a p=5.77e−04 (FIG. 9).
In another example, the role of candidate CSC genes was delineated by evaluation of a 5-gene model (FIG. 10). The genes NRP1, BMI1, JAG1, MSI1, and HTATIP were used in a Kaplan-Meier recurrence plot. The 5-gene model gave separation of triple negative breast cancer patients into the Recurrence and No Recurrence groups with a p=1.04e−03 (FIG. 10). Whereas the 5-gene model exhibited high significance, none of the individual genes demonstrated a p<0.05 individually.

Example 3

Identification of Genes Expressed from Tumorspheres that Increase the Performance of Gene Set Models for Identifying the Recurrence-Free Patient Subset in Breast Cancer

The identification of critical genes from cancer stem cells and tumor initiating cancer cells is exemplified by analysis of in vitro formation of tumorspheres from human cancer cell lines. In this example, tumorspheres were derived from four human breast cancer cell lines: MDA-MB-231, SUM-159, MCF7, and Hs578T. Notably, two of these cell lines are derived from breast cancer patients with basal-like morphology. The method illustrates a means to determine the relative expression of candidate cancer stem cell and tumor initiating cancer cell genes by comparing tumorspheres with two-dimensional cell culture. The fold-change in gene expression was measured by Q-PCR for RNA samples isolated from Tumorspheres or from two-dimensional cell culture of human breast cancer cell lines. The genes KRT5, SERPINF1, S100A4, RGL1, FAP, TGFBR3, FBLN5, MLPH, FADS2, TFPI, PPAP2B, SEPT9, CYBRD1, UGDH, TRAM2, PCDH9, and DLC1 were evaluated in this analysis. The plots show the fold change of Tumorsphere/2D culture on a log 2 scale for the three cell lines for several of the genes (FIG. 11). Genes from tumor initiating cancer cell, EMT, and cancer stem cell discriminators (Table 1) that were changed in 3 out of the 4 human breast cancer cell lines between tumorspheres and 2D cultures were: KRT5, MMP1, RGL1, FAP, TGFBR3, FBLN5, STC1, FADS2, IGFBP4, VIM, IBGFBP3, UGDH, ENPP2, IER3, SNAIL2, MYL9, DOCK10, DAB2, ROR1, FERMT2, MAP1B, PVRL3, PTGER4, CDH2, CTGF, and TAGLN. Genes that were upregulated or downregulated in a consistent manner between all four of the human breast cancer cells lines between tumorspheres and 2D culture were: CYBRD1, DLC1, MLPH, MYL1, PCDH9, S100A4, SERPINF1, TFPI, and TRAM2. The genes KRT5, SERPINF1, S100A4, RGL1, FAP, TGFBR3, FBLN5, MLPH, FADS2, TFPI, PPAP2B, SEPT9, CYBRD1, and UGDH were determined to be upregulated. Likewise, the genes TRAM2, PCDH9, and DLC1 were downregulated. There were several genes that were upregulated >2 fold in tumorspheres of all 3 cell lines compared to 2D, including: KRT5, SERPINF1, S100A4, RGL1, FAP, TGFBR3, FBLN5, PPAP2B, SEPT9, CYBRD1 and UGDH. Genes that were upregulated in tumorspheres of all 3 cell lines compared to 2D (at least changed in the same direction): MLPH, FADS2, TFPI, DLC1, TRAM2 and PCDH9. Similarly, with genes from tumor initiating property discriminators (Table 1) that were changed in 2 out of the 3 human breast cancer cell lines [MCF7, SUM159, Hs578T] were: ARID5B, MALAT1, NT5E, CNR1, HIPK2, FOXO1, NUP188, FXYDS, FKBP5, and DDB2. Also, genes from tumor initiating property discriminators (Table 1) that were changed in all 3 of the human breast cancer cell lines were: BASP1, FKBP5, FLII, POLR2A, PTGS1, SH3BGRL3, and SSBP2.
Evaluation of the role of these genes in predicting breast cancer recurrence was evaluated in triple negative breast cancer (TNBC) patients. In this study, the tumor biopsies of 178 TNBC patients were examined for RNA profiling levels of different genes on the Affymetrics U133 microarray chip. Expression values for genes of interest were computed. A Kaplan-Meier display was outputted for the study, in which patient medical records were used to determine the ‘event-free fraction’ from 0-7 years, for the group of genes [KRT5, SERPINF1, S100A4, RGL1, FAP, TGFBR3, FBLN5, MLPH, FADS2, TFPI, PPAP2B, SEPT9, CYBRD1, UGDH, TRAM2, PCDH9 and DLC1, 16 gene model] and other similar models were determined. In one example, using a 16 gene model from the group [CYBRD1, FAP, TRAM2, PPAP2B, TFPI, PCDH90, RGL1, FADS2, SERPINF1, DLC1, SEPT9, MLPH, S100A4, FBLN5, and TGFBR3], it was determined that these genes fit a model to separate Likely to Recur from Not Likely to Recur patients with high statistical significance, p=5.83e−08 (FIG. 12). The AUC for this separation of patient outcome was determined to be AUC=0.72. A Relative Risk was calculated to be 2.4 with a 95% Confidence Interval range from 1.7-3.4.
In another determination of multigene models, a second 5-gene group consisting of CYBRD1, FAP, PPAP2B, TRAM2 and TFPI genes was examined on the same triple negative breast cancer patient group. A Kaplan-Meier display was outputted for the study where patient medical records were used to determine the ‘event-free fraction’ from 0-7 years. It was determined that these genes fit a model to separate Likely to Recur from Not Likely to recur patients, with a p-value between the ‘Recurrence’ and ‘No recurrence’ subgroups of patients of p=2.19e−08 (FIG. 13). The AUC for this separation of patient outcome was determined to be AUC=0.69 (FIG. 13). A Relative Risk was calculated to be 2.4 with a 95% Confidence Interval range from 1.7-3.4.
In addition, tumorsphere cultures were utilized to screen for the upregulation or downregulation of candidate genes under conditions in which the tumorspheres were treated with chemotherapeutic agents, cytotoxic agents, cytostatic agents and the like. Particular cells may be enhanced or eliminated by these treatments leading to an increased change in expression of key genes in the remaining tumorspheres in culture (secondary tumorspheres). Information about which genes are upregulated or downregulated or unchanged can be informative in the development of clinical diagnostics, and may be useful as drug response biomarkers. For example, sequential passaging of tumorspheres of SUM159 cells led to upregulation of 11 out of 126 CSC-associated genes (FIG. 14). These genes that are changed upon secondary culture of tumorspheres include: CD44, ENPP2, FBLN5, FN1, IGFBP4, PCOLCE, PPAP2B, S100A4, SEMA5A, VCAN, and VIM. Furthermore, expression of upregulated genes was enhanced further in paclitaxel-treated tumorspheres. Other culture conditions such as Matrigel are also relevant to the discrimination of gene sets that show changed regulation under such culture conditions. Correlations observed between the presence of certain expressed genes and specific drug treatment, such as with agents that target cancer stem cells, are expected to change the expression levels of these genes in the drug response.
In an additional refinement, genes were evaluated with regard to differential expression in particular cell types. In this mode, we evaluate the genes that are expressed in mesenchymal (M) tumor cells, and low or not expressed in epithelial (E) tumor cells, and low or not expressed in fibroblast (F) non-tumor cells [M-high, E-low, Fibroblast-low] (FIG. 15). In human tumor specimens, fibroblast or stromal cells are generally not of cancer cell origin, and are not expected to be derived from cancer stem cells. Therefore, screens that exclude the fibroblast or stromal cells, may aid in the definition of cancer stem cells, tumor initiating cancer cells, and mesenchymal associated cancer cells. Examples of M-high, E-low, Fibroblast-low genes include: CDH2, CHN1, CTGF, CYP1B1, DNAJB4, EML1, ENPP2, HAS2, MAP1B, NID1, PDGFC, PRR16, PRRX1, PVRL3, ROR1, SDC2, SNAI1, SNAI2, TNFAIP6, and VCAN. Other genes may have similar properties where different cut-off values will be utilized for discriminating preferential expression between mesenchymal cancer cell and fibroblast-type cells. These genes and gene groups showing differential properties between mesenchymal-type and fibroblast-type cells are considered to describe properties of cancer stem cells, tumor initiating cancer cells, and cancer associated mesenchymal cells in human primary cancer specimens. It is important to determine whether the overexpression and underexpression of genes in this group is able to identify patients that may have a likelihood to recur following chemotherapy treatment, or may have a likelihood to respond and not have evident clinical recurrence after chemotherapy treatment. To evaluate whether the M-High, E-Low, Fibroblast-Low gene subgroup may be sufficient to determine reoccurrence outcome, a cohort of 178 Triple negative breast cancer patient tumor specimens were examined for RNA profiling levels of different genes on the Affymetrics U133 microarray chip. Expression values for genes of interest were computed. A Kaplan-Meier display was outputted for the study where patient medical records were used to determine the ‘event-free fraction’ from 0-7 years. The M-High, E-Low, Fibroblast-Low group of CDH2, CHN1, CTGF, CYP1B1, DNAJB4, EML1, ENPP2, HAS2, MAP1B, NID1, PDGFC, PRR16, PRRX1, PVRL3, ROR1, SDC2, SNAI1, SNAI2, TNFAIP6, and VCAN [19-gene model]), determined that these genes fit a model to separate Likely to Recur (Recurrence) from Not Likely to Recur (No Recurrence) patients with high statistical significance of p=1.9e−03, with an AUC of 0.723 (FIG. 16 A, B). Discrimination of subgroups of these genes with high statistical significance was outputted. This subgroup of the genes that were M-High, E-Low, Fibroblast-Low was derived by drop-one-out iterations with statistical performance assessments, to yield the 6 gene set including genes: CHN1, CTGF, DNAJB4, PRRX1, PVRL3, and VCAN having high statistical significance of a p=4.7e−04 separating the No Recurrence versus Recurrence patients, and with an AUC of 0.71 (FIG. 16 C, D) for the model test.
It is striking that candidate genes derived from either human tumor cell line culture measurements in tumorspheres or from discrimination between mesenchymal and fibroblast gene expression levels and patterns, are both a means to rank and derive multi-gene models that identify patients separating No Recurrence and Recurrence groups.

Example 4

Gene Ranking Based on Gene Expression Variance and Polarity

Genes from the CSC, EMT, and Tumor Initiating origin (Table 1) were evaluated for variance in expression by analyzing the range in expression values amongst a group of samples. The samples used in this evaluation may be from multiple sources, including human cancers, or from models of cancers, such as patient-derived xenografts, or mouse xenografts of human tumor cells, or from comparisons between human cancer cell lines. In the present evaluation, 178 human breast cancer specimens, and associated gene expression microarray data were compared for each gene in the array. The mean expression value and the variance distribution amongst the gene members of the set, relative to all the genes on the array, were processed. In this determination, genes were ranked by increased [cv category A, Table 2], mid-level, or decreased variances. Thus, genes of interest would be determined to have a variance rank (cv.rank), a percentage of variance relative to all genes on the array (percent.cv), and a mean expression, allowing an assessment of relative value. CV category A is indicative of genes that are more variable amongst the sample specimens. CV category B and C indicate genes where the gene expression is not as variable as other genes in the set, and for the specimens tested.
Genes from the CSC, EMT, and Tumor Initiating origin (Table 1) were also inspected for their likelihood to change between increased versus decreased expression, termed Polarity (Table 2). Simulations of 92-143 cycles resulting from 1000 model iterations were processed to output a median.polarity value, a Polarity Percentile, and a median.p (P value of the likelihood of significance based on Highly Significant being a gene that had a consistent Polarity of increased or decreased expression level (Table 2)). In order to discriminate high value of Polarity amongst the genes in the group, an arbitrary cut-off of p<0.01 was established that indicated the genes with <20% or >80% value in Polarity.
Accordingly, subgrouping of significant genes based on Variance and Polarity amongst a large group of human breast cancer specimens, was processed. FIG. 17, shows that the 25 genes that are highly ranked by the Variance criteria, are more variable than all genes in the most Variance group (below the horizontal line, p<0.01), and significantly more variable than the All Genes in the array. FIG. 18, shows that the 25 most highly ranked genes have Polarity values that are consistently of one polarity or the other relative to all the genes in the dataset. Genes that have a Polarity value of 1.00 or near 1.00 have approximately equal probability of being increased or decreased expression when used in a statistical model.
As shown above, selection of Variance and Polarity criteria may be assembled to determine gene ranking prioritization of membership in statistical models for response criteria and clinical outcome data, such as with patient recurrence with the example of human breast cancers. The example was conducted on triple negative breast cancer, but it is not likely to be limited to this cancer type or disease. In the present example, triple negative breast cancer patients (n=178) were evaluated by standard histopathology criteria and determined to be of the triple negative breast cancer subtype by examining ER, PR, and HER2 by immunohistochemistry and pathology of tumor biopsy specimens. Following their diagnosis, patients were treated with standard chemotherapy and their survival was evaluated over the next seven years, so that outcome data was available for the patient cohort. In one test, gene subsets identified by the above rank ordering strategies were evaluated in breast cancer patient specimens with RNA expression data formed from profiling of patient tumors. In addition, recurrence and/or survival data from the patients was utilized.
Genes used in the addition test were at least one from the group of the 25 highest ranked genes based on Variance and Polarity: CYBRD1, INTS8, RGS4, FBXO21, PRRX1, POLS, ID3, OLFML3, PRSS16, CIRBP, CHN1, SERPINE1, SH2B3, ZBTB16, TWIST1, EIF3S9, DPF2, CDH11, CTGF, CCNB1, VIL2, UBE2S, APLP2, MGP, and NOL8. However, more genes may populate these lists based on the criteria of Variance and/or Polarity. Alternatively, genes may be mixed based on Good Ranking Status as one criteria (Table 2) and on Appearance in Tumorsphere cultures as an independent criteria.
In one execution of statistical models, the 25 highest ranked genes based on Variance and Polarity [CYBRD1, INTS8, RGS4, FBXO21, PRRX1, POLS, ID3, OLFML3, PRSS16, CIRBP, CHN1, SERPINE1, SH2B3, ZBTB16, TWIST1, EIF3S9, DPF2, CDH11, CTGF, CCNB1, VIL2, UBE2S, APLP2, MGP, and NOL8] were iteratively sampled by logistic regression and leave-one-out sampling to identify the 18 top ranked genes. The top 18 ranked genes (CYBRD1, INTS8, RGS4, FBXO21, PRRX1, POLS, ID3, OLFML3, PRSS16, CIRBP, CHN1, SERPINE1, SH2B3, ZBTB16, TWIST1, EIF3S9, DPF2, CDH11, CTGF, CCNB1, VIL2, and UBE2S) were highly ranked, where the models containing these genes showed a p<0.05 significance for 13 of the genes. When analyzed in Kaplan-Meier Time to Recurrence plots, the top 18 gene model (multigene model) showed a separation of triple negative breast cancer patients into the Recurrence and No Recurrence groups with a p<0e+00 (FIG. 19). In comparison, with the 25 gene model, the AUC computed in Receiver Operator Curve analysis 0.92.
Extension of the analysis to reduce the number of genes in the model, and to discriminate the driver genes in the model was also conducted with these gene lists. Starting from the top 18 genes determined by Variance, Polarity, and Leave-one-out, statistical output in Kaplan-Meier Recurrence plots determined that a refinement to a 12-gene model (CYBRD1, INTS8, RGS4, FBXO21, PRRX1, ID3, CHN1, SERPINE1, ZBTB16, TWIST1, CCNB1, UBE2S) had a highly significant statistical value for the separation of triple negative breast cancer patients into the Recurrence and No Recurrence groups with a p<0e+00 (FIG. 20). In comparison, for this 12 gene model, the computed AUC was 0.894 in Receiver Operator Curve analysis. In the top 12 gene model [CYBRD1, INTS8, RGS4, FBXO21, PRRX1, ID3, CHN1, SERPINE1, ZBTB16, TWIST1, CCNB1, UBE2S], all 12 genes had a p<0.05.
Likewise, starting from the top 12 genes by Variance, Polarity, and Leave-one-out statistical output in a Kaplan-Meier Recurrence plot was executed. It was determined that a refinement to give a 6-gene model (CYBRD1, RGS4, PRRX1, ID3, SERPINE1, and ZBTB16) had a highly significant statistical value for the separation of triple negative breast cancer patients into the Recurrence and No Recurrence groups with a p<1.22e−10 was observed (FIG. 21). In comparison, for this 6 gene model, the AUC was computed to 0.816 in Receiver Operator Curve analysis. In the top 6 gene model [CYBRD1, RGS4, PRRX1, ID3, SERPINE1, and ZBTB16], all 6 genes had a p<0.05. Genes that are either upregulated or downregulated may contribute to these models.
Genes that were candidate genes based on laboratory Q-PCR measurements, such as with Tumorsphere culture analysis were also co-evaluated in gene ranking strategies based on Variance and Polarity. In one example, the four genes CYBRD1, SERPINF1, FAP, and PPAP2B, individually indicated that increased expression is found in tumorsphere cultures, possibly possessing increased levels of cancer stem cells, and statistical output in Kaplan-Meier Recurrence plots were executed. Further refinements also gave several multi-gene models that had highly significant statistical value for the separation of triple negative breast cancer patients into the Recurrence and No Recurrence groups. For example, CYBRD1, SERPINF1, FAP, PPAP2B with RGS4 and PRRX1 (Tumorsphere 6-gene model) yielded a p=1.21e−07, and an AUC of 0.756 (FIG. 22). In comparison, an 8-gene model combining CYBRD1, SERPINF1, FAP, PPAP2B with RGS4, PRRX1, FBXO21, and ID3 was a highly significant model with a p=2.66e−12, and the computed AUC of 0.821 in Receiver Operator Curve analysis. Genes that are either upregulated or downregulated may contribute to these models. In addition, the weighting of each gene may be individually determined.
These examples demonstrate that combinations of genes may be fit to combinatorial models defining an ability to separate patients into No Recurrence and Recurrence groups. Definition of these patient subgroups may contribute to and represent functionally relevant subgroups for future treatments with agents that inhibit or kill cancer stem cell, tumor initiating cancer cell, and tumor associated mesenchymal.

TABLE 2

Rank Order of Genes based on Variance and Polarity

							cv	times used
		median.	cv.	cv.	percent.	Mean.	cate-	in a	polarity,	polarity,
gene	median.p	polarity	rank	percentile	cv	expression	gory	simulation	percentile	category

CYBRD1	0.000447844	0.923472345	11663	87.41567981	10.0269784	5.971931781	A	110	1%	Good
INTS8	0.000616548	1.046942757	10715	80.31029831	8.527228238	7.72922699	A	118	99%	Good
RGS4	0.000822959	1.039597077	9718	72.83765552	7.507724639	5.633178734	A	109	99%	Good
FBXO21	0.002553602	0.955077734	9377	70.28181682	7.189143411	6.666660827	A	98	7%	Good
PRRX1	0.002666276	0.955583314	12330	92.4149303	11.78954373	5.545155189	A	115	8%	Good
POLS	0.004302918	1.040256137	9156	68.62539349	7.022201602	8.08865953	A	110	99%	Good
ID3	0.004354395	1.051773745	12848	96.29740669	14.70610253	5.535638216	A	105	100%	Good
OLFML3	0.006954545	0.940273485	12594	94.39364413	12.96726543	7.370975636	A	110	3%	Good
PRSS16	0.007902452	0.966097766	11165	83.68310598	9.159501564	5.330501554	A	101	12%	Good
CIRBP	0.008174492	0.937708742	11088	83.10598111	9.041599393	8.081095769	A	127	2%	Good
CHN1	0.008563674	1.022740484	9985	74.83885474	7.757662112	6.898214366	A	125	93%	Good
SERPINE1	0.008883279	1.036139646	10308	77.25978114	8.086722676	5.571009865	A	120	97%	Good
SH2B3	0.009050152	1.015900731	9231	69.18752811	7.078451596	6.474525203	A	109	87%	Good
ZBTB16	0.010047024	0.94722274	12784	95.81771848	14.1791896	5.075007435	A	96	4%	Good
TWIST1	0.011391626	1.075579454	13014	97.54159796	16.25944106	4.434120542	A	131	100%	Good
EIF3S9	0.012128966	1.022312183	7411	55.54639484	6.001613725	8.702829032	A	107	92%	Good
DPF2	0.013138673	0.964324655	8899	66.69914556	6.840009078	7.097320284	A	137	11%	Good
CDH11	0.01348098	0.947875852	13115	98.29860591	17.80084339	6.042363365	A	110	4%	Good
CTGF	0.013752874	0.891138612	13252	99.32543846	21.7169806	6.984676613	A	99	0%	Good
CCNB1	0.013821791	1.03419766	12498	93.67411183	12.4850567	6.668403951	A	143	96%	Good
VIL2	0.014892512	1.021687837	10633	79.6956978	8.428163329	7.216439561	A	122	92%	Good
UBE2S	0.01528562	0.979664602	12199	91.43306851	11.39222508	8.019233778	A	120	22%	Good
APLP2	0.015343287	0.945443626	11216	84.06535752	9.245491905	8.232277003	A	122	3%	Good
MGP	0.015353658	0.897058445	13037	97.71398591	16.50926949	11.06831798	A	116	0%	Good
NOL8	0.015537274	1.029697179	9964	74.68145705	7.740372192	6.31811628	A	116	95%	Good
IGFBP7	0.015696722	1.001137554	13261	99.39289462	22.23554394	7.560811784	A	97	58%	Poor
CTNNB1	0.015988802	0.951844112	10396	77.91935242	8.16482316	8.201877814	A	123	6%	Good
CLDN4	0.017338142	1.027368568	11703	87.71548493	10.13444062	7.266147398	A	120	95%	Good
LUM	0.018167638	0.969794331	13334	99.94003897	31.39303237	6.718396361	A	109	15%	Good
POSTN	0.018770994	0.996917249	13150	98.56093539	18.36687176	5.726984481	A	100	50%	Poor
MYL9	0.019542106	0.912226471	13172	98.72582821	18.91298463	6.971630847	A	91	1%	Good
MARCH8	0.021280706	1.045476492	12267	91.94273722	11.60879333	6.13962269	A	116	99%	Good
SFPQ	0.022377897	0.979004471	8483	63.58117224	6.580693249	7.083770203	A	92	21%	Good
RBM15	0.022727478	0.976622098	9464	70.93389297	7.268488277	6.931840351	A	105	19%	Good
ERBB3	0.022850985	0.973024232	8807	66.00959376	6.778733253	6.14068606	A	131	16%	Good
BTG1	0.023096581	0.950500449	10829	81.16474292	8.672832814	8.570409728	A	114	5%	Good
TPD52	0.023234514	1.010603283	12791	95.87018438	14.23442693	7.516610524	A	114	81%	Good
NUP62	0.023300327	0.979867486	7366	55.20911408	5.982017633	6.942777818	A	107	22%	Good
FAP	0.023506772	1.004061302	12595	94.40113926	12.96841883	6.024875876	A	98	66%	Poor
PPAP2B	0.023748859	0.958007438	11497	86.17148853	9.732243317	7.364726161	A	117	8%	Good
DNAPTP6	0.023803226	1.019898578	10924	81.87678009	8.826829556	8.065916216	A	109	91%	Good
FYN	0.023827838	1.011799028	10351	77.58207165	8.123534234	5.285960467	A	116	82%	Good
LRP2	0.024274966	1.047306842	11282	84.56003598	9.350029818	5.10881537	A	113	100%	Good
PTGS1	0.024541016	1.036329331	8941	67.01394094	6.86973762	6.488618599	A	101	97%	Good
NID2	0.026681166	1.004849338	12654	94.84335182	13.34147014	5.638125179	A	118	67%	Good
TUBB3	0.026780898	1.015886784	10213	76.54774397	7.969793643	8.770726503	A	112	87%	Good
GSK3B	0.026815921	0.972999591	10647	79.80062959	8.445282837	6.67339815	A	117	16%	Good
TGFB3	0.027153519	0.988004808	7602	56.97796432	6.111650196	7.149941128	A	117	30%	Good
ECHDC2	0.027187608	0.962803694	10435	78.21166242	8.201884038	7.930889688	A	112	10%	Good
CD97	0.027551005	1.034406195	10260	76.90001499	8.027118571	6.814423675	A	120	97%	Good
PSORS1C2	0.027621367	1.039509647	8787	65.8596912	6.764158437	5.918339127	A	132	98%	Good
SEMA5A	0.028511613	0.992572064	8588	64.3681607	6.645242779	6.353350689	A	124	40%	Poor
ANXA5	0.028615878	0.968071623	7480	56.06355869	6.043746331	9.219588653	A	117	13%	Good
DUSP10	0.028825844	1.016882409	11222	84.11032829	9.25433409	5.724554818	A	107	88%	Good
SPARC	0.02885106	0.995086707	13194	98.89072103	19.51614725	8.08817232	A	117	46%	Poor
ITGBL1	0.03009402	0.97201951	11693	87.64053365	10.12036996	4.757603281	A	127	16%	Good
PROCR	0.030248086	0.98139105	9946	74.54654475	7.727404282	3.756769316	A	111	23%	Good
SH3BGRL	0.030545781	0.961674856	11894	89.14705441	10.54424222	7.827599607	A	100	10%	Good
STARD13	0.030604916	0.982192308	7706	57.75745765	6.158437666	6.393370904	A	112	25%	Good
TGFB1I1	0.030610931	0.950910009	12834	96.19247489	14.57446646	5.700066466	A	106	5%	Good
CCDC92	0.030971772	0.974551506	7063	52.93809024	5.845490151	7.344843621	A	113	17%	Good
REEP5	0.032462419	0.961120433	11061	82.90361265	9.011099714	8.221658311	A	103	9%	Good
SKIV2L2	0.033171044	0.971929622	8915	66.81906761	6.849853258	7.005727204	A	103	16%	Good
CTBS	0.033180484	1.009276888	8499	63.70109429	6.590587498	5.022598712	A	123	78%	Good
CD59	0.033380327	0.945253168	11724	87.87288263	10.17727689	9.163708042	A	125	3%	Good
DCN	0.034121792	0.939694	13254	99.34042872	21.89477494	7.775674163	A	110	2%	Good
CITED2	0.0343109	0.94685154	12543	94.01139259	12.69664229	5.516936887	A	106	4%	Good
FN1	0.034487776	1.016406804	12870	96.46229951	14.92352135	9.972283396	A	123	88%	Good
SERPINF1	0.034497864	0.953997594	13007	97.48913206	16.23023863	7.832252688	A	128	7%	Good
IGFBP3	0.034699744	1.039429523	12801	95.94513566	14.3166115	7.569712229	A	125	98%	Good
SPOCK1	0.035214101	1.009987543	11581	86.8010793	9.883514348	6.096964305	A	98	79%	Good
MYO10	0.035286043	1.024906164	11025	82.63378804	8.962687791	6.388851103	A	103	94%	Good
SSBP2	0.035521187	0.961467549	9918	74.33668116	7.703550364	5.518643423	A	133	9%	Good
NIFUN	0.035619214	0.966546046	9340	70.00449708	7.162211196	8.349568156	A	109	12%	Good
LOC388397	0.0364212	0.981205093	8983	67.32873632	6.894703408	4.703300964	A	106	23%	Good
PVRL3	0.036718216	1.019618758	7223	54.13731075	5.919449686	4.987661462	A	139	91%	Good
FSTL1	0.036811565	0.98276311	12923	96.8595413	15.30596317	8.82415883	A	107	25%	Good
CLDN3	0.03719767	0.963887258	12945	97.02443412	15.53709116	7.186124553	A	119	11%	Good
PSMD8	0.037260823	1.01636378	9559	71.64593015	7.353649877	9.122068456	A	117	87%	Good
AGPS	0.037341896	1.037880541	11082	83.06101034	9.035022529	5.401390365	A	113	98%	Good
IL13RA1	0.037527615	0.998913896	11398	85.42947084	9.551024598	6.550059063	A	122	54%	Poor
DAB2	0.037570858	0.971910665	11779	88.28511468	10.30606776	6.667650435	A	99	15%	Good
GNG11	0.037926758	1.001886527	12878	96.52226053	14.99439355	5.427606628	A	121	61%	Poor
FKBP5	0.038677717	0.939940183	12957	97.11437566	15.64621134	5.197440247	A	99	3%	Good
SLC38A2	0.03907335	0.963817018	10171	76.23294858	7.92910891	8.820154703	A	109	11%	Good
UGDH	0.039881238	0.974540813	11701	87.70049468	10.133079	6.283186837	A	132	17%	Good
DBR1	0.040269416	0.995654341	7764	58.19217509	6.184331265	4.51888702	A	113	47%	Poor
SNRPN	0.040441891	0.950901014	12178	91.27567081	11.31125892	7.586200142	A	126	5%	Good
LDHA	0.040556405	0.996967123	7059	52.90810973	5.84445503	11.06438928	A	123	50%	Poor
FLNB	0.041865103	1.007390443	9479	71.04631989	7.285309575	7.657875997	A	105	74%	Good
KLF13	0.042315813	0.988769976	9971	74.73392295	7.743982203	5.548923778	A	119	32%	Good
KDELR2	0.043186242	0.981348028	10563	79.17103882	8.360291047	8.528602961	A	104	23%	Good
TMED5	0.043392451	1.007080236	9532	71.44356168	7.325146241	6.67780651	A	117	73%	Good
TGIF	0.043625131	1.006912658	11045	82.7836906	8.984191126	6.772798007	A	109	72%	Good
CHPT1	0.044294713	0.952973286	11711	87.77544596	10.15630195	7.869692753	A	109	6%	Good
ETS1	0.044353829	1.018617495	7734	57.96732124	6.171734411	5.639135859	A	116	89%	Good
SSR1	0.044677648	0.976786917	10065	75.438465	7.8232335	7.239957828	A	102	19%	Good
DDIT4	0.04481242	1.039489206	12520	93.83900465	12.58433447	8.085099068	A	106	98%	Good
ZEB1	0.044974662	0.995883789	9042	67.77094888	6.936086814	4.55858065	A	97	47%	Poor
CHST2	0.045018767	0.957579565	12713	95.28556438	13.68689954	4.804627223	A	125	8%	Good
TUBB	0.04517446	1.000077573	7484	56.0935392	6.044508267	10.61181493	A	118	56%	Poor
GREM1	0.045477406	0.989901322	13088	98.09623745	17.18917409	4.548108995	A	108	34%	Poor
ARHGAP24	0.046706069	1.023356549	7716	57.83240893	6.162376786	4.750632859	A	109	93%	Good
LOC283824	0.047299197	0.974994664	9943	74.52405936	7.725810909	5.674670787	A	122	18%	Good
ID4	0.047333161	0.926290837	13175	98.7483136	19.04594117	5.36801418	A	116	1%	Good
CALD1	0.047335546	0.975672609	11349	85.06220956	9.469529157	5.752658554	A	111	18%	Good
SETD5	0.047365706	0.97984763	8565	64.19577275	6.634601298	7.613185373	A	130	22%	Good
BGN	0.048339531	0.997665346	12113	90.78848748	11.12728011	6.262085281	A	111	52%	Poor
HSPA2	0.048826033	0.931279326	13013	97.53410283	16.25384777	5.74573716	A	105	2%	Good
PREPL	0.048920223	0.968369698	9528	71.41358117	7.3227757	6.227496378	A	112	14%	Good
PCOLCE	0.049394724	0.989680586	12569	94.20626593	12.80969501	6.161098888	A	111	34%	Poor
H2AFZ	0.049792598	0.985456902	8993	67.4036876	6.900564544	9.620431347	A	118	27%	Good
FTL	0.050542822	0.982057704	7698	57.69749663	6.152683998	11.24754452	A	115	24%	Good
MMP2	0.050599232	0.977082837	12576	94.25873182	12.86264339	7.851665299	A	116	19%	Good
TNFAIP6	0.050693153	1.037703169	12844	96.26742617	14.6630354	5.769113086	A	117	97%	Good
FBN1	0.050864318	0.964885169	12917	96.81457053	15.24599852	6.382559387	A	132	11%	Good
CDKN2C	0.051563247	0.971522551	9958	74.63648628	7.737459293	5.54430355	A	122	15%	Good
TGFBR3	0.05190111	0.953558514	13144	98.51596462	18.28297346	5.651828325	A	115	7%	Good
TACSTD1	0.052113157	0.973656827	12362	92.6547744	11.90521366	8.666395185	A	130	17%	Good
APOBEC3G	0.052130686	0.995981903	7937	59.48883226	6.27897091	6.518417958	A	110	47%	Poor
MLPH	0.052329976	0.990255975	12512	93.77904362	12.53458876	8.238630904	A	142	35%	Poor
THY1	0.053261819	0.991225808	11726	87.88787288	10.17882942	6.873596074	A	122	38%	Poor
PTGER4	0.053909735	0.976041605	11357	85.12217059	9.487759113	5.55424783	A	120	19%	Good
CD44	0.054299143	1.018693434	12061	90.39874082	10.95846408	7.958652143	A	116	89%	Good
RYBP	0.054526411	0.982445682	6847	51.31914256	5.753487942	7.151997263	A	106	25%	Good
HIF1A	0.054595768	0.991093187	11460	85.89416879	9.669752382	8.316753302	A	119	37%	Poor
PLOD2	0.054711258	0.978663474	12893	96.63468745	15.14814977	6.535608639	A	99	21%	Good
CD24	0.05479293	1.031812692	12081	90.54864338	11.03343396	9.645361683	A	118	96%	Good
MAPT	0.054829639	0.986985328	10640	79.74816369	8.434202522	6.175932422	A	102	29%	Good
ZMYM2	0.054890756	0.977908822	8984	67.33623145	6.894733811	4.045907178	A	111	20%	Good
FKSG49	0.054912365	1.019174242	7435	55.72627792	6.017473205	8.740764899	A	94	89%	Good
KLF10	0.055936512	1.009127749	11685	87.58057263	10.08895166	6.739359769	A	119	77%	Good
SMYD3	0.056458642	1.012037521	12029	90.15889672	10.87096281	7.021742714	A	107	82%	Good
PGK1	0.056867134	0.992071554	10570	79.22350472	8.364592863	8.9091619	A	130	39%	Poor
NAT10	0.057027928	0.994938582	8072	60.50067456	6.341315286	7.937177411	A	124	45%	Poor
TNS3	0.057440585	0.960931473	12219	91.58297107	11.43623106	7.526211843	A	113	9%	Good
ALG5	0.057593273	0.99255466	10540	78.99865088	8.324688197	7.87469492	A	118	40%	Poor
CXCL2	0.05793095	1.013077618	12926	96.88202668	15.3566365	5.060339355	A	116	84%	Good
EMP3	0.058019782	0.989955188	11904	89.2220057	10.56790767	7.564953976	A	118	35%	Poor
METTL7A	0.058125645	1.000924279	8913	66.80407735	6.847593838	6.032787886	A	108	58%	Poor
ECOP	0.058250035	0.988456949	10913	81.79433368	8.807370769	8.156107951	A	120	31%	Good
DNAJB1	0.058387278	0.99463299	9030	67.68100735	6.929318486	7.824524157	A	88	44%	Poor
SEPT6	0.058499125	1.001453977	10447	78.30160396	8.213386956	5.611716712	A	110	60%	Poor
FOXO1A	0.058684039	0.991662529	7269	54.48208664	5.938461181	6.427190252	A	122	39%	Poor
KNTC2	0.05875687	1.019577799	13062	97.90136411	16.83772312	4.876486199	A	113	91%	Good
SDF4	0.058901527	0.996027205	9563	71.67591066	7.355976181	7.244552352	A	119	47%	Poor
LTF	0.058991843	0.891657126	13282	99.55029231	22.88262723	9.078743245	A	111	0%	Good
PMP22	0.059411595	0.953937134	12782	95.80272823	14.17441451	8.430933931	A	115	7%	Good
CEBPD	0.059658661	1.001791786	9140	68.50547144	7.010599231	6.572230051	A	106	61%	Poor
THBS1	0.059998233	0.994531318	11946	89.53680108	10.66998094	5.646315594	A	124	44%	Poor
TGIF2	0.060179816	1.004051263	9055	67.86838555	6.941024143	6.378831142	A	101	65%	Poor
PSMB7	0.061335145	1.01191009	8348	62.56932994	6.48988523	9.130438587	A	118	82%	Good
MMP1	0.062035976	0.922115184	13339	99.97751462	35.83680532	4.419777459	A	116	1%	Good
CYP1B1	0.062435749	0.964387374	13308	99.74516564	24.66200778	6.280096841	A	107	11%	Good
SCGN	0.062736617	1.009032736	7024	52.64578024	5.82868347	6.679948767	A	99	77%	Good
MEF2A	0.062915503	0.99079724	10416	78.06925498	8.180122908	4.507490413	A	118	36%	Poor
NRP1	0.062947993	1.005506589	12642	94.75341028	13.24482768	4.919936938	A	130	69%	Good
MMP9	0.063140539	0.95536991	12743	95.51041823	13.85506057	8.579089506	A	112	8%	Good
PPP3CA	0.063429532	0.968182261	11224	84.12531854	9.25664422	7.268082335	A	121	14%	Good
TGFBI	0.063449576	1.017707534	12162	91.15574876	11.26661692	8.60122753	A	142	88%	Good
ISGF3G	0.063691853	0.99289378	10546	79.04362165	8.332493251	8.601769853	A	127	41%	Poor
STAM	0.063955366	0.993307853	10968	82.20656573	8.885919946	6.485197543	A	137	42%	Poor
CDH1	0.063990642	0.986256849	12438	93.22440414	12.22373315	6.919764517	A	114	28%	Good
SEPT9	0.064233513	0.981691306	9086	68.10073452	6.963536599	6.461530497	A	126	24%	Good
KDELR3	0.064321613	0.989231356	11717	87.82041673	10.1645103	7.313461377	A	100	33%	Good
ARID5B	0.064431404	0.973368765	11337	84.97226803	9.447693266	7.790965233	A	125	17%	Good
RGL1	0.064552908	0.994087901	10265	76.93749063	8.03376252	7.540015464	A	109	44%	Poor
COL1A1	0.064582159	0.993702236	12042	90.25633338	10.91231407	7.001737454	A	117	43%	Poor
ATP2B1	0.065112625	0.991672492	11195	83.90795983	9.212229629	4.081174457	A	102	39%	Poor
CDKN1A	0.065400067	1.012062062	8839	66.24943787	6.800113431	7.792606502	A	102	83%	Good
LRIG1	0.065419254	0.936986999	12494	93.64413131	12.46306291	5.693379708	A	128	2%	Good
CCND2	0.065595281	0.984122713	8107	60.76300405	6.359827224	5.566803935	A	117	26%	Good
MSX1	0.065792082	0.997608989	10048	75.31104782	7.810901502	4.388789279	A	127	51%	Poor
IER3	0.065942671	1.011484853	12218	91.57547594	11.43448028	8.273296205	A	101	81%	Good
FGFR2	0.066022239	1.00281586	10523	78.8712337	8.303116129	5.273177578	A	102	64%	Poor
FRAS1	0.066232058	1.018215725	8104	60.74051866	6.359422969	5.571262561	A	106	89%	Good
BUB1	0.066374098	1.010229466	8094	60.66556738	6.356090093	5.008199211	A	111	80%	Good
TAGLN	0.066753768	0.945502863	13240	99.23549693	21.34313683	8.023282275	A	112	3%	Good
FADS2	0.066894084	1.00928935	12092	90.63108979	11.0644294	7.025023687	A	118	78%	Good
TPM1	0.06769776	0.951736201	12608	94.49857593	13.03768935	8.198073725	A	133	5%	Good
PLAA	0.068144961	1.006889525	7456	55.88367561	6.030267694	4.624008733	A	119	72%	Good
FBLN5	0.068145172	0.966143729	11007	82.49887573	8.94296832	6.828876438	A	124	12%	Good
VIM	0.068301136	0.990511107	9956	74.62149603	7.735132821	11.32438972	A	101	36%	Poor
DPYSL3	0.068327827	0.999031789	12043	90.26382851	10.91821131	6.179769285	A	107	55%	Poor
TP53	0.068890725	1.002054383	12540	93.98890721	12.69380463	5.450920278	A	104	62%	Poor
SH3BGRL3	0.068989975	0.998889705	8519	63.85099685	6.604720209	8.473259078	A	141	54%	Poor
PDGFC	0.069010155	0.96995612	13076	98.00629591	17.04374598	6.827112736	A	126	15%	Good
VCAN	0.069297615	1.000936397	13136	98.4560036	18.17202162	5.291336585	A	111	58%	Poor
IGFBP4	0.069437136	0.968345316	12817	96.06505771	14.42612452	7.456892762	A	107	14%	Good
METTL2B	0.069484582	1.017701034	8334	62.46439814	6.48229014	3.487954209	A	115	88%	Good
BAT3	0.069508605	0.998700011	7285	54.60200869	5.944249708	8.654222218	A	103	54%	Poor
ZBTB38	0.069949149	0.966746157	11310	84.76989957	9.403118607	6.946326518	A	118	13%	Good
EIF4ENIF1	0.070126638	0.989140522	7777	58.28961175	6.191371812	6.104279535	A	127	33%	Good
PPM1D	0.070191618	0.987304421	9155	68.61789837	7.021412099	4.886494829	A	117	29%	Good
TUBA3	0.070205258	0.981819985	11966	89.68670364	10.71903946	9.146891529	A	133	24%	Good
CSTF1	0.070220098	0.991464442	6869	51.48403538	5.762287305	5.4732504	A	114	38%	Poor
ATP13A3	0.070749942	0.993536962	7089	53.13296357	5.856718599	6.019358571	A	124	42%	Poor
STC1	0.070875003	1.026596717	12493	93.63663619	12.45013807	5.828474049	A	117	94%	Good
MYCBP2	0.070973157	0.981188131	11001	82.45390496	8.930354974	7.56106142	A	107	22%	Good
LMCD1	0.070998528	0.959786862	10786	80.84245241	8.610524808	6.885739155	A	117	8%	Good
S100A4	0.071013984	0.98603156	12234	91.69539799	11.49326185	8.514013401	A	115	27%	Good
SDC2	0.071226367	0.949185923	12849	96.30490181	14.70925771	7.304027171	A	121	4%	Good
NUCKS1	0.071342918	0.988673035	7914	59.31644431	6.265676817	8.208101691	A	120	31%	Good
KRT5	0.071586657	0.961563148	13226	99.13056513	20.60166991	8.123189295	A	112	10%	Good
USP22	0.071976141	1.020473168	7115	53.32783691	5.87046935	4.829598306	A	112	91%	Good
MYL1	0.072597769	1.018112974	7693	57.66002099	6.150561576	5.445345463	A	116	89%	Good
FXYD3	0.072830343	0.985294431	10235	76.71263679	8.005600752	8.756347961	A	119	26%	Good
AKAP2	0.072864843	0.986609499	12437	93.21690901	12.22222858	6.19271358	A	129	28%	Good
ENPP2	0.072910324	0.951935449	13190	98.86074052	19.40597917	5.846518734	A	125	6%	Good
MAST4	0.074053136	1.00872382	7510	56.28841253	6.061291271	6.683026188	A	121	76%	Good
LRPAP1	0.07418212	0.988766696	7837	58.73931944	6.223163164	7.410948634	A	102	32%	Good
ECM1	0.074561655	0.992787288	7717	57.83990406	6.164484692	7.046216208	A	112	41%	Poor
STAU1	0.074831662	0.989242699	7908	59.27147354	6.263780625	9.021766442	A	125	33%	Poor
GAS7	0.074852579	0.989957448	8577	64.28571429	6.640121271	5.827292079	A	127	35%	Poor
MAP4K4	0.075044686	0.987534308	9962	74.6664668	7.740300545	5.896665079	A	109	29%	Good
HS3ST2	0.075415011	1.013784175	6960	52.16609204	5.800160915	6.232013975	A	112	85%	Good
PLEKHC1	0.075632005	0.965296409	12368	92.69974517	11.9137684	5.30124757	A	115	12%	Good
NSF	0.076431045	0.988756452	7948	59.57127867	6.283194251	7.482771953	A	118	31%	Good
MT2A	0.076533192	0.978656007	8735	65.46994454	6.728458889	10.93221467	A	96	21%	Good
MMP14	0.076548535	1.006404677	6730	50.44221256	5.703151797	6.774690793	A	115	70%	Good
MLF1	0.076721003	0.999914598	12412	93.0295308	12.12445146	5.966685543	A	108	56%	Poor
NUP37	0.076823725	0.998581101	9775	73.26487783	7.5586067	6.985063829	A	109	53%	Poor
BDH2	0.076885147	0.952537387	11735	87.95532904	10.19544234	6.189184891	A	120	6%	Good
MRPL42	0.077137429	0.988894716	9201	68.96267426	7.053594083	7.812151962	A	94	32%	Good
ETNK1	0.077272351	0.968555234	11794	88.3975416	10.3308613	4.768183348	A	123	14%	Good
C10ORF7	0.077852801	1.006595942	10188	76.36036576	7.944578698	8.478487642	A	149	71%	Good
WEE1	0.077876722	0.988024797	7868	58.97166842	6.238141723	7.164204426	A	117	31%	Good
LTBP2	0.07791878	0.986831097	11283	84.5675311	9.350076099	6.999420183	A	104	28%	Good
KCNMA1	0.077966763	0.993302798	11198	83.93044521	9.218242947	5.152484658	A	138	42%	Poor
SORBS2	0.079306562	0.990643114	9580	71.80332784	7.370629882	6.524991842	A	118	36%	Poor
NR2F1	0.079595536	0.966615968	12086	90.58611902	11.05286316	5.066994165	A	131	13%	Good
DPT	0.079735764	0.952182936	12967	97.18932694	15.71852287	5.047312052	A	107	6%	Good
HNMT	0.080033585	0.99127788	7084	53.09548793	5.855770954	4.780660847	A	101	38%	Poor
DNAJC13	0.080338554	1.002215106	6821	51.12426923	5.739985685	5.195115649	A	105	62%	Poor
NEBL	0.080499686	0.977443415	8240	61.75985609	6.432740239	6.039167608	A	100	20%	Good
KPNA3	0.080518202	1.003541345	7372	55.25408484	5.98392567	7.118066712	A	123	65%	Poor
SCCPDH	0.08111085	0.994540059	10085	75.58836756	7.849323848	6.954962555	A	110	44%	Poor
TBX3	0.081153545	0.96702269	12143	91.01334133	11.20941391	5.639524448	A	109	13%	Good
CDH2	0.081459821	1.009182615	7268	54.47459152	5.937989375	4.968761682	A	111	78%	Good
AMMECR1	0.0823953	1.033372614	12395	92.90211363	12.06524327	6.173349279	A	109	96%	Good
PLP2	0.082539275	0.992919803	10271	76.9824614	8.041833925	8.615654634	A	107	41%	Poor
HS2ST1	0.082735053	1.015298178	9067	67.95832709	6.949328432	4.213077405	A	118	86%	Good
CNIH4	0.082912768	1.019177709	12626	94.63348823	13.1654349	6.813580509	A	132	90%	Good
KIAA1217	0.083305849	1.015571208	6802	50.98186179	5.731883811	7.811091686	A	120	86%	Good
EIF2C2	0.08349556	1.034393949	10900	81.69689702	8.780506716	6.065294901	A	118	96%	Good
ICMT	0.084397089	1.002920288	8879	66.54924299	6.825259531	6.307082341	A	107	64%	Poor
IER5	0.085452591	1.005831521	12704	95.21810823	13.62496933	6.878858514	A	125	69%	Good
NOTCH2	0.085543756	0.974260289	8875	66.51926248	6.823958309	7.514383233	A	109	17%	Good
VTCN1	0.08578649	0.981477259	12921	96.84455104	15.29110333	8.054099209	A	91	23%	Good
MAFF	0.086497949	1.007013768	12479	93.53170439	12.39351987	6.009550547	A	114	73%	Good
SNAI2	0.086574535	0.962420986	13201	98.94318693	19.74765063	5.322643176	A	117	10%	Good
FMNL2	0.086841007	1.028262012	7678	57.54759406	6.144369796	5.447286508	A	122	95%	Good
WFDC2	0.087257206	0.975271754	12388	92.84964773	12.02598283	7.118478554	A	109	18%	Good
GABARAPL1	0.087277933	0.995266597	8892	66.64667966	6.833603312	5.847708059	A	124	46%	Poor
CASP8	0.087421397	1.002688822	7957	59.63873482	6.286765402	5.897421355	A	104	64%	Poor
NID1	0.087843474	0.989749576	12306	92.23504722	11.72130736	5.171256868	A	110	34%	Poor
RAB23	0.087857858	0.992586751	9113	68.30310298	6.984974883	4.712004618	A	121	40%	Poor
STC2	0.087904728	0.985726133	10262	76.91500525	8.031336724	6.919758563	A	116	27%	Good
AIM1	0.088211291	0.981017219	12633	94.68595413	13.18773458	8.094941948	A	108	22%	Good
ZBTB20	0.088478936	0.976149944	9230	69.18003298	7.078161877	6.102837084	A	120	19%	Good
PAK2	0.088737701	1.001933025	11298	84.67995803	9.381673543	5.839209371	A	124	62%	Poor
ANK3	0.088801077	0.987388726	11407	85.496927	9.566173955	6.157982418	A	122	29%	Good
WNT5A	0.088962096	0.99843273	11659	87.3856993	10.01262977	4.690399636	A	113	53%	Poor
EXT1	0.089319348	1.008762726	7389	55.38150202	5.990970898	6.381146541	A	119	76%	Good
COL11A2	0.091309029	1.010180563	6725	50.40473692	5.700008942	6.460264931	A	105	80%	Good
RIN2	0.091892713	0.989724047	11680	87.54309699	10.0710586	6.867399743	A	118	34%	Poor
SGK3	0.092100841	0.986226616	13078	98.02128616	17.08613324	4.076982724	A	120	28%	Good
CEP350	0.092399536	1.005254166	9349	70.07195323	7.168109756	6.272724389	A	115	68%	Good
BASP1	0.093953473	1.004659362	12424	93.11947234	12.17698969	8.634079152	A	126	67%	Good
STK39	0.094477437	0.977826945	12255	91.85279568	11.55444502	5.290319946	A	106	20%	Good
PTX3	0.094693519	0.961407318	13323	99.85759256	28.1324859	4.311307552	A	88	9%	Good
NFYA	0.095713033	0.99611783	7144	53.54519562	5.882806875	6.129571257	A	110	47%	Poor
MME	0.096026621	0.996843604	12051	90.32378954	10.94090406	6.208853699	A	109	49%	Poor
SPP1	0.0973073	1.034622206	13089	98.10373257	17.20035112	9.602476381	A	108	97%	Good
CPS1	0.097351825	0.984522832	12708	95.24808874	13.65547073	4.654868227	A	120	26%	Good
TFPI	0.097929983	0.999249541	12427	93.14195773	12.18453708	3.95334446	A	115	55%	Poor
ATIC	0.098124163	0.992285019	9002	67.47114376	6.907580925	8.193399336	A	88	40%	Poor
PTGFR	0.098956076	0.990940671	7852	58.85174636	6.231009874	3.58087155	A	111	37%	Poor
TOB2	0.099371499	0.997273579	8416	63.07899865	6.540253029	6.04147893	A	113	50%	Poor
MARS	0.100664359	0.991546405	11440	85.74426623	9.638801634	6.697500891	A	111	39%	Poor
SRP54	0.101071913	0.977955491	9320	69.85459451	7.14508165	7.668295957	A	102	21%	Good
KRT6B	0.101564496	1.004845522	13059	97.87887873	16.77676281	7.477978183	A	94	67%	Good
EPPK1	0.103045521	0.997754807	10739	80.49018138	8.553905214	7.057086919	A	112	52%	Poor
CES1	0.103365221	0.993725109	11159	83.63813521	9.152249574	5.861298358	A	103	43%	Poor
THEM2	0.104189298	0.997303091	10852	81.33713086	8.714432399	7.038182753	A	122	51%	Poor
PFKFB3	0.104988685	1.005322731	11482	86.05906161	9.710066398	6.755897972	A	100	69%	Good
RAD23B	0.104990727	0.997097047	10412	78.03927447	8.175735873	7.563156654	A	111	50%	Poor
EMP1	0.107749455	0.989455201	13242	99.25048718	21.3926789	6.131865312	A	107	33%	Poor
MMP7	0.110763102	1.00661322	13280	99.53530205	22.77381948	8.379755002	A	121	71%	Good
PILRB	0.113239479	0.994640009	11425	85.6318393	9.6085534	6.967198571	A	125	45%	Poor
LAMA2	0.113712178	0.986215791	10475	78.51146755	8.257917231	4.37876049	A	112	28%	Good
CHST11	0.11401461	1.006465565	10070	75.47594064	7.833199439	5.091184862	A	114	71%	Good
ANXA6	0.116727976	0.987943215	10398	77.93434268	8.167392935	7.920064873	A	107	30%	Good
GTF3C3	0.116876154	0.992607052	9233	69.20251836	7.079273961	6.423745468	A	96	41%	Poor
LTBP1	0.118087056	1.001437177	9031	67.68850247	6.931433998	7.06272816	A	127	59%	Poor
PTGER2	0.11986875	0.997094826	7944	59.54129816	6.282635374	4.598143852	A	125	50%	Poor
FBLN1	0.121359663	0.998315961	8781	65.81472043	6.759868092	6.842070183	A	102	53%	Poor
GPRC5A	0.122961107	0.987611101	12765	95.67531105	14.05034413	7.555993599	A	101	30%	Good
GNPDA1	0.12509288	0.999533044	8302	62.22455404	6.467482305	6.879013896	A	99	56%	Poor
SWAP70	0.130474933	0.981547602	10117	75.82821166	7.875858511	5.830347266	A	111	24%	Good
ATP1B3	0.131830106	0.999420865	10091	75.63333833	7.851367399	9.545326018	A	111	56%	Poor
MMP10	0.164779488	0.998396792	10920	81.84679958	8.822068421	3.745111984	A	112	53%	Poor
JTV1	0.003204972	1.022919703	5826	43.6666167	5.352863101	8.174106486	B	109	93%	Good
GBX2	0.014449505	1.026339068	5505	41.26068056	5.226970939	4.051663642	B	115	94%	Good
XPNPEP1	0.015396931	0.968931722	6297	47.19682207	5.529644192	6.996324653	B	131	14%	Good
SNED1	0.018307564	1.013080486	4187	31.38210163	4.813588383	4.998283657	B	109	84%	Good
SLC11A1	0.018929015	1.02313728	4569	34.24524059	4.927450102	6.763001666	B	127	93%	Good
ADAMTS7	0.023034398	1.014112283	6444	48.29860591	5.583706838	5.138618655	B	128	85%	Good
GTPBP1	0.023807917	1.020920722	3745	28.06925498	4.672224091	5.919511762	B	114	92%	Good
TRAM2	0.023969061	0.977420929	5905	44.25873182	5.380820292	6.604805949	B	100	20%	Good
MAPK14	0.02547542	1.014325417	4002	29.99550292	4.758622153	6.580294539	B	116	85%	Good
LRP1	0.027488558	0.991611677	3340	25.03372808	4.543243403	6.835485128	B	131	39%	Poor
GNB2L1	0.030890249	0.974775467	6459	48.41103283	5.590550504	7.910934413	B	110	18%	Good
CAMK2B	0.031161237	0.993300817	5411	40.55613851	5.195039495	4.696629449	B	137	42%	Poor
TGFB1	0.032505161	0.996191955	5423	40.64608005	5.199865271	6.279307709	B	112	48%	Poor
TRIO	0.035568563	1.019459509	3437	25.76075551	4.571591941	5.632634989	B	129	90%	Good
CLIC5	0.038302723	1.026880935	6620	49.61774846	5.653526997	4.250555849	B	126	95%	Good
DNAJB4	0.040587612	1.008282713	6322	47.38420027	5.538953258	4.353922559	B	118	75%	Good
TMEM63A	0.042033336	1.008461021	5167	38.72732724	5.121087294	6.129186684	B	113	75%	Good
PSD	0.042758143	1.01174101	6652	49.85759256	5.666413442	5.045674777	B	104	81%	Good
NT5E	0.042867461	1.008493845	4214	31.58447009	4.821287185	5.740755289	B	100	75%	Good
PSMA5	0.042932042	1.001177544	6455	48.38105232	5.5883386	8.466188705	B	112	59%	Poor
FBXL18	0.043121041	1.023814184	5681	42.57982311	5.285119356	4.842451458	B	100	94%	Good
ST3GAL5	0.043414465	0.986198492	6259	46.9120072	5.514777943	7.189554894	B	120	27%	Good
SMPD1	0.046288268	1.021137066	4337	32.50637086	4.861915365	5.313541576	B	110	92%	Good
ARF3	0.04663004	0.98395595	3521	26.39034627	4.598461631	8.526647644	B	117	25%	Good
GAPDH	0.047951792	0.996452041	3875	29.04362165	4.719251752	12.43369353	B	131	48%	Poor
COPZ2	0.048545006	1.002960187	6255	46.88202668	5.512841087	6.965871158	B	109	65%	Poor
IL8RA	0.048654499	1.011601337	4331	32.46140009	4.860430883	7.247323975	B	125	81%	Good
ELL2	0.052024721	1.008268505	4768	35.7367711	4.985786697	5.312578136	B	97	75%	Good
FXYD5	0.052604063	1.021814942	6562	49.18303103	5.631635597	7.298651887	B	101	92%	Good
KLK3	0.053451536	1.004810946	3937	29.50831959	4.735273566	6.352365244	B	115	67%	Good
ATXN3	0.053853417	1.010645532	4884	36.60620597	5.028065588	3.94858219	B	103	81%	Good
C1ORF63	0.054714725	0.991147236	4311	32.31149753	4.853094096	6.43854867	B	122	37%	Poor
ELP4	0.056871199	1.008878238	5812	43.5616849	5.347421718	6.392940346	B	117	77%	Good
PCTK2	0.057264782	0.996619794	3355	25.146155	4.547917914	5.257992978	B	97	49%	Poor
CNOT4	0.058563281	1.015498212	4737	35.50442213	4.975482415	4.647202978	B	113	86%	Good
ARPC5	0.058812533	0.993132789	4151	31.11227702	4.802391759	9.551114774	B	113	42%	Poor
TNRC6B	0.061585503	1.00493343	5387	40.37625543	5.187061915	5.876065572	B	121	68%	Good
CNR1	0.062502761	1.010340114	4953	37.12336981	5.053719929	5.27782066	B	104	80%	Good
WWOX	0.064188007	1.011831147	3392	25.42347474	4.558316791	5.91322382	B	130	82%	Good
PRKCA	0.064519713	1.006996779	5970	44.74591516	5.40503885	5.681840748	B	120	73%	Good
TGFBR1	0.065287493	1.004114214	5849	43.83900465	5.358698653	4.826721378	B	115	66%	Good
UGCGL1	0.066585663	1.019522416	5700	42.72223055	5.2904279	5.572911872	B	112	90%	Good
COPB2	0.068075935	0.993990059	4101	30.73752061	4.787062704	6.1498873	B	128	44%	Poor
TAS2R14	0.068194207	1.002110981	5716	42.8421526	5.299596368	5.423281781	B	104	62%	Poor
DOCK10	0.068227546	1.000096802	6347	47.57157847	5.548950174	3.932605508	B	125	57%	Poor
DCUN1D4	0.069000046	0.99480297	4209	31.54699445	4.820055146	5.788236954	B	118	45%	Poor
FAM53C	0.070065685	1.0040693	3476	26.05306551	4.581495102	7.588488115	B	117	66%	Poor
ZFHX1B	0.071091597	0.997750729	5177	38.80227852	5.122745904	3.463636081	B	112	52%	Poor
PTPRA	0.071162198	0.987805878	4534	33.98291111	4.918221295	5.554080282	B	122	30%	Good
LPIN2	0.071184707	1.005147909	4593	34.42512367	4.936699448	6.303011207	B	122	68%	Good
NDEL1	0.071868675	1.001613164	6610	49.54279718	5.649116738	6.838990595	B	103	60%	Poor
DNM1	0.072039805	1.009705227	3367	25.23609654	4.552789571	6.022019507	B	122	79%	Good
BIN1	0.072858777	0.990182196	6170	46.24494079	5.478892673	7.159294163	B	116	35%	Poor
CREB3L1	0.073155202	1.007507766	4035	30.24284215	4.767435118	6.463534704	B	126	74%	Good
HAS2	0.073188069	1.002887889	5527	41.42557338	5.234724161	4.000166384	B	93	64%	Poor
SDHAL1	0.076750617	1.005968065	3422	25.64832859	4.567198875	8.763855971	B	108	70%	Good
ROR1	0.078740429	1.002352518	4871	36.5087693	5.023136838	4.997233037	B	108	63%	Poor
EIF4E2	0.081254067	1.000733692	6614	49.57277769	5.651590085	6.66540388	B	131	58%	Poor
EML1	0.081877176	0.995097599	5730	42.9470844	5.307363622	3.983998361	B	105	46%	Poor
FAM49B	0.08353526	1.01236775	4153	31.12726728	4.802850912	5.016285835	B	110	83%	Good
CHST7	0.08568526	1.00863996	5293	39.67171339	5.158784112	5.447210971	B	119	76%	Good
QKI	0.087219371	0.990747708	5687	42.62479388	5.28662618	5.984907217	B	113	36%	Poor
SNX6	0.087819189	0.993569263	4075	30.54264728	4.778104354	7.363871414	B	119	43%	Poor
LARP1	0.088542647	0.999517496	3500	26.23294858	4.589809462	8.103120611	B	126	56%	Poor
DDB2	0.09261684	0.983955802	6265	46.95697796	5.517217091	6.18229142	B	95	25%	Good
DDR2	0.098470039	0.988229502	5993	44.9183031	5.411531384	6.215293751	B	129	31%	Good
NOTCH3	0.099498625	1.00410418	6263	46.94198771	5.516037017	7.152488578	B	124	66%	Good
KLF7	0.101220269	1.008573894	5140	38.52495878	5.110202821	7.472285062	B	97	76%	Good
DHRS4	0.106248327	0.98468674	6451	48.3510718	5.586789265	7.014512199	B	127	26%	Good
ERBB4	0.109124088	1.000687663	3438	25.76825064	4.572291059	4.524602834	B	97	57%	Poor
DNMT3A	0.109264487	1.006092904	3791	28.41403088	4.687603161	6.163119538	B	102	70%	Good
PCNX	0.113247075	1.001876769	3414	25.58836756	4.565391125	4.147407073	B	121	61%	Poor
ZCCHC6	0.114181724	1.001416528	6669	49.98500974	5.671624773	7.650567691	B	104	59%	Poor
CSNK2A1	0.120905109	1.000904796	5796	43.44176285	5.338501705	7.355664723	B	120	58%	Poor
GGCX	0.132200055	0.996237684	4727	35.42947084	4.973062314	6.097073153	B	111	48%	Poor
RNF2	0.138291474	1.000404524	3994	29.9355419	4.755613151	4.75417006	B	113	57%	Poor
BICD1	0.000997899	0.995479784	845	6.333383301	3.596604619	7.276725939	C	121	46%	Poor
POU2F2	0.015878559	1.019510995	1713	12.83915455	4.000005069	5.555314232	C	121	90%	Good
CG018	0.021754796	0.988878543	3195	23.94693449	4.491712915	5.011038491	C	111	32%	Good
ACTG1	0.027652893	0.990805209	817	6.123519712	3.579911364	12.49096002	C	112	37%	Poor
SLC25A16	0.031872768	1.024251496	1673	12.53934942	3.98374567	4.458671727	C	117	94%	Good
SYT11	0.039319097	0.997453577	3100	23.23489732	4.463242088	4.819834744	C	116	51%	Poor
FSTL4	0.04042613	1.015843697	2236	16.75910658	4.180153387	5.647671942	C	103	87%	Good
BCL2	0.042220431	0.991340249	2757	20.66406836	4.358849031	6.059490629	C	105	38%	Poor
NFRKB	0.045890333	1.010114992	532	3.987408185	3.384743692	6.644913041	C	116	80%	Good
POLR2A	0.046934535	1.009962241	2162	16.2044671	4.151777952	4.480060266	C	121	79%	Good
ERN1	0.048382257	1.007297966	2398	17.97331734	4.236720625	6.364227225	C	120	73%	Good
KLHL20	0.048456385	1.012940488	918	6.880527657	3.639701372	5.337923827	C	106	84%	Good
C7ORF25	0.049043508	1.012484816	408	3.058012292	3.287209075	4.84821256	C	128	83%	Good
ADRBK1	0.050711378	1.008428438	2625	19.67471144	4.310653823	7.010186885	C	111	75%	Good
LARS	0.052978046	0.989025076	2533	18.98515965	4.282906087	8.118859388	C	108	33%	Good
MAP1B	0.055360088	1.009544692	2542	19.0526158	4.287262187	4.653352601	C	109	78%	Good
KIAA0194	0.055451709	1.000342672	228	1.708889222	3.097201468	7.701227206	C	111	57%	Poor
LUZP1	0.057217627	1.002400713	14	0.104931794	2.393743761	7.491755018	C	120	63%	Poor
ALKBH1	0.058662383	0.998142886	2570	19.26247939	4.293919654	7.377745876	C	119	53%	Poor
NUP188	0.059353633	1.012540825	1558	11.67740968	3.937055029	5.178103055	C	113	83%	Good
SNAI1	0.060508864	1.012521793	1283	9.616249438	3.815857079	6.683897367	C	124	83%	Good
HIPK2	0.063008144	1.001895285	2778	20.82146605	4.366234811	6.265548604	C	119	61%	Poor
FLJ43663	0.064537856	1.007741024	201	1.506520762	3.048786797	3.814065669	C	124	74%	Good
PCDH9	0.0653874	1.014570227	1091	8.17718483	3.729993631	4.277113912	C	108	86%	Good
RERE	0.065776079	1.006397817	552	4.137310748	3.40503803	4.361013509	C	102	70%	Good
PER1	0.068415183	1.002393399	104	0.779493329	2.872860021	6.980244659	C	125	63%	Poor
MR1	0.072032842	0.996628802	1185	8.881726878	3.773163213	5.982442371	C	96	49%	Poor
C11ORF17	0.073505038	0.990299378	2821	21.14375656	4.379304256	5.673302422	C	129	36%	Poor
HPS1	0.073548841	1.013649511	149	1.116774097	2.976950121	6.476592219	C	105	85%	Good
SCNM1	0.074244986	0.996396531	1547	11.59496327	3.932748595	9.004397408	C	128	48%	Poor
NDST2	0.07429773	1.008817643	769	5.76375356	3.547817017	6.939415527	C	123	77%	Good
RAD51L1	0.074460851	0.997754631	1579	11.83480738	3.944954476	4.097040759	C	129	52%	Poor
CACNA1G	0.075211702	0.997563526	1384	10.37325738	3.858756224	4.800607103	C	101	51%	Poor
EIF1	0.076536456	0.994648703	2108	15.79973018	4.133156616	10.48885468	C	114	45%	Poor
CENTD2	0.080564655	1.006845736	717	5.374006896	3.519616268	6.93084978	C	116	72%	Good
PDE8A	0.08097957	1.005686521	1365	10.23084995	3.85119531	5.819030061	C	122	69%	Good
DVL3	0.081543688	0.998622456	3126	23.42977065	4.469027263	6.455380783	C	124	54%	Poor
DYM	0.081565656	1.009860818	2787	20.8889222	4.371258687	3.964215145	C	125	79%	Good
PDE4A	0.083335146	1.013153044	3266	24.47908859	4.520067813	4.826309346	C	120	84%	Good
FGFR1	0.083804982	1.015614485	2506	18.78279119	4.274916322	6.861838821	C	124	86%	Good
PHF15	0.085726566	1.001440553	2157	16.16699146	4.150070456	6.182042197	C	98	59%	Poor
ST3GAL2	0.085927127	1.001725725	577	4.324688952	3.42893581	5.64176634	C	98	60%	Poor
FLII	0.086013521	0.993969465	1932	14.48058762	4.072676222	7.079464198	C	117	43%	Poor
KLF12	0.08724397	1.006763617	970	7.270274322	3.672972828	5.523407118	C	107	72%	Good
IFNAR1	0.090763099	1.005154032	2673	20.03447759	4.330093106	5.620071998	C	123	68%	Good
PNPLA2	0.092560293	1.001734028	1016	7.615050217	3.69580935	8.421514347	C	113	60%	Poor
ZNF335	0.093172698	1.005406575	624	4.676959976	3.462618042	6.272871357	C	103	69%	Good
ABR	0.094812705	0.999088198	1834	13.74606506	4.040402717	6.62483202	C	107	55%	Poor
CHRD	0.096586451	1.006811688	3162	23.69959526	4.481484519	5.478358766	C	132	72%	Good
CLIC1	0.099728132	1.003507577	2984	22.36546245	4.42832086	9.648849067	C	124	65%	Poor
CLTC	0.100536086	1.007783183	2185	16.37685504	4.159691969	3.48823819	C	135	74%	Good
HCFC1	0.104103183	1.002632451	30	0.224853845	2.602159713	7.280074735	C	109	63%	Poor
PDE4DIP	0.104560793	1.006505039	1654	12.39694199	3.973761821	4.353246344	C	111	71%	Good
IRF5	0.113490731	1.004678089	770	5.771248688	3.548102358	6.202174462	C	118	67%	Good
DLC1	0.114091838	1.002775945	797	5.973617149	3.568438577	5.096913541	C	107	64%	Poor
ARHGEF2	0.117069343	1.001833226	2400	17.9883076	4.236806804	7.531437586	C	116	61%	Poor
KIAA0146	0.123537953	0.996667145	1456	10.91290661	3.892494472	5.304924979	C	94	49%	Poor
RNF8	0.125410979	1.009623546	2831	21.21870784	4.381387893	5.797011142	C	122	78%	Good
HOM_TES_103	0.065328707	0.999101055	#N/A	#N/A	#N/A	#N/A	#N/A	124	55%	Poor

EQUIVALENTS

Those skilled in the art will recognize, or be able to ascertain using no more than routine experimentation, many equivalents to the specific embodiments described herein. Such equivalents are intended to be encompassed by the following claims.

Claims

What is claimed:

1. A method of evaluating or treating a subject, comprising:

a) optionally, acquiring a subject sample;

b) acquiring a value or values that is a function of the level of gene expression for (each of) a plurality of genes selected from a first and/or second and/or third and/or fourth and or/fifth and/or sixth set of genes, wherein

(i) said first set of genes comprises or consists of:

S100A4, SDC2, CYP1B1, FERMT2, CYBRD1, SNAI2, CD44, BIN1, PMP22, CTGF, IGFBP3, ANXA6, MAP1B, TAGLN, ENPP2, FBLN5, PCOLCE, FN1, VIM, PPAP2B, SEM5A, VCAN, NID2, NID1, HAS2, PRRX1, TNFAIP6, CLDN4, FADS2, DDIT4, GNG11, PER1, ISCU, KPNA3, SLC44A1, FKBP5, NR2F1, MLPH, DOCK10, RGS4, IGFBP4; and

(ii) said second set of genes comprises or consists of:

S100A4, SDC2, CYP1B1, FERMT2, CYBRD1, SNAI2, CD44, BIN1, ENPP2, FBLN5, PCOLCE, FN1, VIM, PPAP2B, SEM5A, VCAN, CLDN4, FADS2, DDIT4, GNG11, PER1, ISCU, KPNA3, SLC44A1, FKBP5, IGFBP4; and

(iii) said third set of genes comprises or consists of:

(iv) said fourth set of genes comprises or consists of:

(v) said fifth set of genes comprises or consists of:

FERMT2, BIN1, PMP22, CHN1, CTGF, IGFBP3, NID2, CLDN4; and

(vi) said sixth set of genes comprises or consists of:

HAS2, BIN1, PCOLCE, FERMT2, CTGF, IGFBP3, NID2, SLC44A1, FKBP5, and MLPH; and

(c) responsive to said value or values:

(i) classifying said subject; or

(ii) administering treatment comprising an agent that inhibits or kills cancer associated mesenchymal cells, tumor initiating cancer cells, or cancer stem cells, to said subject; provided that, if (c)(ii) is not performed the acquisition in (a) or (b) comprises directly acquiring; thereby evaluating or treating said subject.

2-3. (canceled)

4. The method of claim 1, wherein c(i) and c(ii) are performed.

5-11. (canceled)

12. The method of claim 1, wherein step b) said plurality comprises, or consists of, a first gene and a second gene.

13. The method of claim 12, wherein step b) comprises acquiring a value that is a function of the level of gene expression of said first gene and the level of gene expression of said second gene.

14-17. (canceled)

18. The method of claim 13, wherein step b) comprises acquiring a first value that is a function of the level of gene expression of two or more genes of said plurality and a second value that is a function of the level of gene expression of one of the genes of the plurality.

19. The method of claim 1, further comprising acquiring a value or values that is a function of the level of gene expression of a gene not listed in claim 1.

20. (canceled)

21. The method of claim 13, wherein step b) comprises acquiring a value that is a function of the level of gene expression of said first gene, the level of gene expression of said second gene, and a weighting factor.

22. (canceled)

23. The method of claim 18, wherein said first value is a function of a first weighting factor and said second value is a function of a second weighting factor.

24. (canceled)

25. The method of claim 1, wherein said value or values is a function of a comparison with a reference criterion.

26. The method of claim 25, wherein said value or values is further a function of the determination of whether the level of gene expression has a preselected relationship with a reference criterion.

27. (canceled)

28. The method of claim 1, further comprising determining if said value or values has a preselected relationship with a reference criterion.

29-31. (canceled)

32. The method of claim 1, wherein said subject sample is a tumor sample, a tissue sample, a biopsy, blood sample, plasma sample, serum sample, interstitial fluid sample.

33. (canceled)

34. The method of claim 1, wherein a first value or values is acquired for a first location in said subject sample.

35. The method of claim 34, wherein a second value or values is acquired for a second location in said subject sample.

36. (canceled)

37. The method of claim 35, comprising determining if said first value or values and/or said second value or values has a preselected relationship with a reference criterion.

38-42. (canceled)

43. The method of claim 34, wherein said first value or values is a function of the level of gene expression of each gene of a plurality of genes at said first location.

44. (canceled)

45. The method of claim 1, comprising, responsive to the acquisition of said value or values one or more of:

(1) stratifying a patient population; and/or

(2) identifying or selecting the subject as likely or unlikely to respond positively to a treatment; and/or

(3) selecting a treatment; and/or

(4) prognosticating the time course of the disease in the subject and/or

(5) measuring the response at the end of therapy and predicting the long term outcome; and/or

(6) determining the cancer stem cell population as a predictor of response to a treatment or therapy.

46-61. (canceled)

62. A method of assaying in a subject sample the level of gene expression product of a plurality of genes selected from a first and/or second and/or third and/or fourth and/or fifth and/or sixth set of genes, wherein:

(i) said first set of genes comprises or consists of:

(ii) said second set of genes comprises or consists of:

(iii) said third set of genes comprises or consists of:

(iv) said fourth set of genes comprises or consists of:

(v) said fifth set of genes comprises or consists of:

FERMT2, BIN1, PMP22, CHN1, CTGF, IGFBP3, NID2, CLDN4; and

(vi) said sixth set of genes comprises or consists of:

HAS2, BIN1, PCOLCE, FERMT2, CTGF, IGFBP3, NID2, SLC44A1, FKBP5, and MLPH; comprising a first agent capable of interacting with a gene expression product of a plurality of genes selected from a first and/or second and/or third and/or fourth and/or fifth and/or sixth set of genes; and wherein the method comprises assaying the level of gene expression product of the plurality of genes.

63-77. (canceled)

78. A reaction mixture comprising:

a plurality of detection reagents; and

a plurality of target nucleic acid molecules derived from a subject, wherein each of the plurality of detection reagents comprises a plurality probes to measure the level of gene expression product of a plurality of genes selected from a first and/or second and/or third and/or fourth and/or fifth and/or sixth set of genes, wherein:

(i) said first set of genes comprises or consists of:

(ii) said second set of genes comprises or consists of:

(iii) said third set of genes comprises or consists of:

(iv) said fourth set of genes comprises or consists of:

(v) said fifth set of genes comprises or consists of:

FERMT2, BIN1, PMP22, CHN1, CTGF, IGFBP3, NID2, CLDN4; and

(vi) said sixth set of genes comprises or consists of:

HAS2, BIN1, PCOLCE, FERMT2, CTGF, IGFBP3, NID2, SLC44A1, FKBP5, and MLPH.

79-110. (canceled)